Production of bacteriocin-like inhibitory substances (BLIS) by Streptococcus salivarius strains isolated from the tongue and throat of children with and without sore throat

Cepas de Streptococcus salivarius, isoladas de crianças com e sem dor de garganta, foram testadas quanto à produção de bacteriocina contra Streptococcus pyogenes. Os resultados mostraram que as crianças que não tinham dor de garganta possuiam, na boca, cepas de bactérias produtoras de substâncias inibidoras semelhantes à bacteriocina contra S. pyogenes.

Bacteriocin-like substances production by Streptococcus-salivarius against oral bacteria

Samples of tongue and bucal mucosa surfaces were obtained from six healthy subjects with the purpose of isolating S. salivarius. It was verified that 47 out of 48 S. salivarius strains produced bacteriocin-like substances against at least one of the indicator species: Actinomyces viscosus, Rothia dentocariosa, Streptococcus pyogenes, Staphylococcus aureus, Streptococcus mutans and Streptococcus sanguis. The method employed to test for bacteriocin production was that of deferred antagonism. The results showed that there was a high antagonism against R. dentocariosa, S. pyogenes and A. viscosus; extremely low against S. mutans and S. sanguis and no inhibition for S. aureus.

Äidinmaito - terveysjuomaa ja normaalibakteereita

The chemical composition of breast milk has been studied in detail in the past decades. Hundreds of new antibacterial and antiviral components have been found. Several molecules have been found to promote the proper function of neonatal intestine. However, microbiological studies of breast milk have been, until recently, focused mainly on detecting harmful and pathogenic bacteria and viruses. Natural microbial diversity of human milk has not been widely studied before the work reported in this thesis. This is mainly because breast milk has traditionally been thought to be sterile - even if a certain amount of commensal bacteria have usually been detected in milk samples. The first part of this licentiate thesis contains a short literature review about the anatomy and physiology of breast feeding, human milk chemical and microbiological composition, mastitis, intestinal flora and bacteriocins. The second part reports on the experiments of the licentiate work, concentrating on the microbial diversity in the milk of healthy breast-feeding mothers, and the ability of these bacteria to produce antibacterial substances against pathogenic bacteria. The results indicate that human milk is a source of commensal bacteria for infant intestine. 509 random isolates from 40 breast milk samples were isolated and identified by 16S rRNA sequencing. Median bacterial count was about 600 colony forming units per milliliter. Over half of the isolates were staphylococci, and almost one third streptococci. The most common species were skin bacteria Staphylococcus epidermidis and oral bacteria Streptococcus salivarius and Streptococcus mitis. Lactic acid bacteria, identified as members of Lactobacillus-, Lactococcus- and Leuconostoc -genera, were found in five milk samples. Enterococci were found in three samples. A novel finding in this study is the capability of these commensal bacteria to inhibit the growth of pathogens. In 90 precent of the milk samples commensal bacteria inhibiting the growth of Staphylococcus aureus were found. In 40 precent of samples the colonies could block the growth completely. One fifth of the isolated Staph. epidermidis strains, half of Str. salivarius strains, and all lactic acid bacteria and enterococci could inhibit or block the growth of Staph. aureus. In further study also Listeria innocua- and Micrococcus luteus active isolates were found in 33 and 11 precent of milk samples (out of 140). Furthermore, two Lactococcus lactis isolates from the breast milk were shown to produce bacteriocin nisin, which is an antimicrobial molecule used as a food preservative. The importance of these human milk commensal bacteria in the development of newborn intestinal flora and immune system, as well as in preventing maternal breast infections, should be further explored.

A new type of Brazilian propolis: Prenylated benzophenones in propolis from Amazon and effects against cariogenic bacteria

Ethanol extracts of four propolis samples (E1-E4) from Manaus (Brazilian Amazon) were analysed by HPLC/DAD/ESI-MS/MS and GC/EIMS. The major constituents of E2 and E4 were analysed by NMR ((1)H and (13)C) and ESI/MS/MS. The main constituents of E2 and E4 are polyprenylated benzophenones: 7-epi-nemorosone, 7-epi-clusianone (major E4 constituents), xanthochymol and gambogenone (major E2 constituents), making up a chemical profile so far unreported for Brazilian propolis. Aristhophenone, methyl insigninone, 18-ethyloxy-17-hydroxy-17,18-dihydroscrobiculatone B, and derivatives of dimethyl weddellianone A and B, propolones, and a scrobiculatone derivative, were detected as minor constituents. Triterpenoids (beta-amyrins, beta-amyrenone, lupeol and lupenone) were ubiquitous and predominant in El and E3. The extracts E2 and E4 were highly active against the cariogenic bacteria Streptococcus mitis, Streptococcus mutans and Streptococcus salivarius. E2 was more active than E4, probably due to a higher content of 2-epi-nemorosone, while the latter was richer in di-hydroxylated compounds. (C) 2010 Elsevier Ltd. All rights reserved.

CARACTERÍSTICAS FÍSICAS E QUÍMICAS DE BEBIDAS LÁCTEAS FERMENTADAS E PREPARADAS COM SORO DE QUEIJO MINAS FRESCAL

Nesta pesquisa procurou-se verificar as características físicas e químicas de bebidas lácteas preparadas com três concentrações de soro de queijo Minas Frescal (30, 40 e 50%), empregando-se dois tipos de culturas lácticas: uma tradicional para iogurte (YC-180) contendo cepas mistas de Streptococcus salivarus subsp. thermophilus, Lactobacillus delbrueckii subsp. lactis e Lactobacillus delbrueckii subsp. bulgaricus e outra (ABY-1) contendo cepas mistas de Lactobacillus delbrueckii subsp. bulgaricus, Lactobacillus acidophillus, Bifidobacteria e Streptococcus salivarius subsp. thermophilus. Constatou-se que as bebidas lácteas apresentaram diferença estatística no tempo zero para os teores de gordura e de extrato seco. À medida em que se elevou a proporção de soro em relação ao leite, os teores de gordura e de extrato seco diminuíram. O teor de proteína também diminuiu à medida em que se aumentou o teor de soro nas bebidas lácteas, embora a diferença não tenha sido tão acentuada quanto as observadas para os teores de gordura e de extrato seco. em relação à lactose, não se constatou diferença entre os tratamentos. Os teores de soro não influenciaram o índice de proteólise das bebidas lácteas. Verificou-se todavia que as bebidas elaboradas com a cultura probiótica ABY-1 apresentaram valores superiores para proteólise quando comparadas às bebidas elaboradas com as culturas YC-180. As bebidas lácteas elaboradas com 30% de soro apresentaram maiores valores para viscosidade. As bebidas elaboradas com a cultura YC-180 apresentaram valores superiores para viscosidade durante o período de armazenamento.

Preparation and antimicrobial activity of gelatin microparticles containing propolis against oral pathogens

Gelatin microparticles containing propolis ethanolic extractive solution were prepared by spray-drying technique. Particle,, with regular morphology, mean diameter ranging of 2.27 mu m to 2.48 mu m, and good entrapment efficiency for propolis were obtained. The in vitro antimicrobial activity of microparticles was evaluated against microorganisms of oral importance (Enterococcus faecalis, Streptococcus salivarius, Streptococcus sanguinis, Streptococcus mitis, Streptococcus mutans, Streptococcus sobrinus, Candida albicans, and Lactobacillus casei). The utilized techniques were diffusion in agar and determination of minimum inhibitory concentration. The choice of the method to evaluate the antimicrobial activity of microparticles showed be very important. The microparticles displayed activity against all tested strains of similar way to the propolis, showing greater activity against the strains of E. salivarius, S. sanguinis, S. mitis, and C albicans.

Antimicrobial activity of the synthetic peptide Lys-a1 against oral streptococci

The peptide LYS-[TRP6]-Hy-A1 (Lys-a1) is a synthetic derivative of the peptide Hy-A1, initially isolated from the frog species Hypsiboas albopunctatus. According to previous research, it is a molecule with broad antimicrobial activity. The objective of this study was to evaluate the antimicrobial activity of the synthetic peptide Lys-a1 (KIFGAIWPLALGALKNLIK- NH2) on the planktonic and biofilm growth of oral bacteria. The methods used to evaluate antimicrobial activity include the following: determination of the minimum inhibitory concentration (MIC) and minimum bactericidal concentration (MBC) in microtiter plates for growth in suspension and quantification of biomass by crystal violet staining and counting of colony forming units for biofilm growth. The microorganisms Streptococcus oralis, Streptococcus sanguinis, Streptococcus parasanguinis, Streptococcus salivarius, Streptococcus mutans and Streptococcus sobrinus were grown in Brain Heart Infusion broth at 37 °C under atmospheric pressure with 10% CO2. The peptide was solubilized in 0.1% acetic acid (v/v) at various concentrations (500-1.9 μg mL-1). Chlorhexidine gluconate 0.12% was used as the positive control, and BHI culture medium was used as the negative control. The tested peptide demonstrated a remarkable antimicrobial effect, inhibiting the planktonic and biofilm growth of all strains tested, even at low concentrations. Thus, the peptide Lys-a1 is an important source for potential antimicrobial agents, especially for the control and prevention of microbial biofilms, which is one of the most important factors in cariogenic processes. © 2012 Elsevier Inc.

Utilização de diferentes culturas lácteas na fabricação da mussarela de leite de búfula

Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES)

„The plug and the bug“ – a case of coronary embolism in bacterial endocarditis

Systemic embolism is a classic complication of infective endocarditis. Coronary involvement and acute myocardial infarction (MI) are rare and increase mortality significantly. Recognising this unusual entity is crucial to provide adequate care. Percutaneous coronary intervention and thrombus aspiration is preferred to thrombolysis, which classically increases intracerebral hemorrhage risk. The present article describes the case of an acute inferior ST-elevated MI due to a Streptococcus salivarius endocarditis in a patient with known bicuspid aortic valve.

Estudo comparativo da microflora bacteriana de doentes portadores de próteses totais acrílicas mucosuportadas e implantosuportadas

Dissertação para obtenção do grau de Mestre no Instituto Superior de Ciências da Saúde Egas Moniz

Population structure and characterization of viridans group streptococci (VGS) including Streptococcus pneumoniae isolated from adult patients with cystic fibrosis (CF)

A study was undertaken to examine the population structure of viridans group streptococci (VGS) in the sputum of adult patients with cystic fibrosis (CF). Freshly expectorated sputa (n=58) from 45 adult CF patients were examined by selective conventional culture on Mitis-Salivarius agar and yielded 190 isolates of VGS. Sequence analyses of the rpnB and 16-23S rRNA ITS genes identified these isolates to belong to 12 species of VGS and included S. anginosus, S. australis, S. cristatus, S. gordonii, S. infantis, S. mitis, S. mutans, S. oralis, S. parasanguinis, S. pneumoniae, S. salivarius and S. sanguinis. The most frequently VGS organism isolated was S. salivarius (47/190; 24.7%), followed by S. mitts (36/190; 19%), S. sanguinis (25/190; 13.2%), S. oralis (20/190; 11.0%), S. pneumoniae (19/190; 10.0%), S. parasanguinis (16/190; 8.4%), S. infantis (11/190; 5.8%), S. gordonii (7/190; 3.7%), S. anginosus (4/190; 2.1%), S. cristatus (2/190; 1.1%), S. australis (1/190; 0.5%), S. mutans (1/190; 0.5%) and S. agalactiae (1/190; 0.5%). All, but four, patients harboured at least one VGS species, which ranged from one to five streptococcal species, with a mean of 2.85 species per patient. There was no clonality at the subspecies level employing ERIC RAPD PCR. Antibiotic susceptibility was determined by Minimum Inhibitory Concentration (MIC) testing against penicillin, erythromycin and ciprofloxacin. Overall, resistance to penicillin with all VGS was 73/190 (38.4%) and 167/190 (87.9%) for erythromycin. With regard to ciprofloxacin, 27/190 (14.2%) were fully resistant, whilst a further 21/190 (11.1%) showed intermediate resistance, which equated to approximately three quarters (74.7%) of isolates being fully sensitive to this agent. In addition, as a comparator control population, we examined antibiotic susceptibility, as above, in a non-CF population comprising 12 individuals (50 VGS isolates), who were not receiving chronic antibiotics. In comparison, 8% and 38% of VGS isolates from non-CF individuals were resistant by disk susceptibility testing to penicillin and erythromycin, respectively. None of the non-CF VGS organisms were resistant to ciprofloxacin, but 42% showed intermediate resistance. (C) 2010 European Cystic Fibrosis Society. Published by Elsevier B.V. All rights reserved.

Longitudinal study of the influence of removable partial denture and chemical control on the levels of Streptococcus mutans in saliva

Some studies have evaluated the salivary levels of mutans streptococci (MS) in removable partial denture (RPD) users. Saliva samples (2.0 mL) were obtained from 31 patients in six periods: (T0): immediately before installation of RPD; (T8): 8 days after T0; (T48): 48 days after T0; (T92): 92 days after T0; (T140): 140 days after T0 and (T189): 189 days after T0. The samples were vortexed and serially diluted from 10(-1) to 10(-6) in 0.05 m phosphate buffer (pH 7.4). From each dilution, 0.025 mL was plated on Mitis Salivarius Bacitracin (MSB). The plates were incubated in 5% CO2 at 37 degreesC for 72 h. There was an increase (t -test, P < 0.05) in the number of MS between periods T0 and T48 (mean/s.d., CFU mL(-1) of saliva): T0: 2.26/4.43 x 10(6) and T48: 0.47/1.48 x 10(8) . After this, intensive treatment with CHX was accomplished in 29 patients. Saliva samples were obtained after treatment in four periods: (T24 h): 24 h after T0; (T14): 14 days after T24 h; (T28): 28 days after T24 h, and (T63): 63 days after T24 h. The number of MS in saliva did not decrease (t -test, P > 0.05). A new CHX formulation was applied in 15 patients. Saliva samples were obtained in periods: (T0): before new CHX application; (T24 h): 24 h after T0 and (T82): 82 days after T0. The new CHX reduced MS levels in saliva: (mean/s.d., CFU mL(-1) of saliva): T0: 6.64/8.47 x 10(6) and T24 h: 3.2/4.27 x 10(5) (sign rank, P < 0.05). In conclusion, there was a significant increase in the number of MS in saliva after the installation of RPD. The intensive treatment with a properly formulated CHX was effective in the reduction of MS, between 24 h and 82 days after its application.

IgA anti-Streptococcus mutans em crianças com e sem cárie dentária

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)

Production of monoclonal antibodies against Streptococcus mutans antigens

Several studies have been conducted in the last decades aiming to obtain an anti-canies vaccine, however some studies have demonstrated cross reactivity between Streptococcus mutans surface antigens and the human cardiac tissue. In this work, the reactivity of five anti-Streptococcus mutans monoclonal antibodies (MoAb) (24A, 56G, G8, E8 and F6) was tested against oral streptococci, cardiac antigens and skeletal and cardiac myosins, aiming to evaluate the specificity of these MoAb. The hybrid producers of immunoglobulins of the IgG 2b class were cloned by limit dilution and expanded in vivo. MoAb were tested by ELISA. The hybrid 24A reacted with S. mutans CCT 1910, S. salivarius CCT 0365 and S. pyogenes T23. No reactivity difference was observed among the tested species. Cross reactivity with heart and cardiac myosin was not confirmed and only reaction with myosin of skeletal muscle was observed (p = 0.0381). The hybrid 56G reacted with all the tested microorganisms and there was statistically significant difference between S. mutans and S. pyogenes T23 (p < 0.001). This hybrid also reacted with myosin of skeletal muscle (p = 0.0095). C8, E8 and F6 presented low reactivity against oral streptococci strains and no reactivity against cardiac antigens. The data of this study showed that the 24A and 56G anti-S. mutans MoAb presented reactivity with S. pyogenes and S. salivarius, reinforcing the occurrence of common antigens between these species. The tested MoAb presented low cross-reactivity with myosin of skeletal muscle, but anti-heart activity could not be confirmed.

Effect of an alcoholic diet on dental caries and on Streptococcus of the mutans group: Study in rats

The objective of this study was to evaluate the effects of an alcohol diet on Streptococcus of the mutans group and on dental caries in the oral cavity of rats. Forty animals were divided into 3 groups according to the following liquid diets: 20% ethanol solution (Alcohol Group, AG), 27% sucrose solution (Isocaloric Group, IG), and water (Control Group, CG). After 56 days, samples were collected and plated on Mitis Salivarius Bacitracin agar to assess the number of colony forming units (CFU/mL) of Streptococcus of the mutans group. The animals were sacrificed and the jaws were removed in order to assess the occurrence of dental caries on the smooth and occlusal surfaces using stereomicroscopy. The data were submitted to ANOVA and Tukey test. The average numbers of CFU/mL (10 3) were: 8.17 (AG), 9.78 (IG), and 5.63 (CG). There was no significant difference among the groups for the occurrence of occlusal caries. Regarding smooth surface caries, in the upper jaw, the caries number in the IG (1.58) was similar to that in the AG (2.06) and in the CG (1.14), and the number of caries in the AG was higher than in the CG; in the lower jaw there was significant difference among the 3 groups: AG (1.14), IG (2.00) and CG (0.43). The diets with the alcohol and sucrose solutions presented a tendency of increasing the colonization by Streptococcus of the mutans group and of increasing the occurrence of smooth surface dental caries in rat molars when compared to the control diet.