Microbial Evaluation Of Traumatized Teeth Treated With Triple Antibiotic Paste Or Calcium Hydroxide With 2% Chlorhexidine Gel In Pulp Revascularization.

Revascularization outcome depends on microbial elimination because apical repair will not happen in the presence of infected tissues. This study evaluated the microbial composition of traumatized immature teeth and assessed their reduction during different stages of the revascularization procedures performed with 2 intracanal medicaments. Fifteen patients (7-17 years old) with immature teeth were submitted to the revascularization procedures; they were divided into 2 groups according to the intracanal medicament used: TAP group (n = 7), medicated with a triple antibiotic paste, and CHP group (n = 8), dressed with calcium hydroxide + 2% chlorhexidine gel. Samples were taken before any treatment (S1), after irrigation with 6% NaOCl (S2), after irrigation with 2% chlorhexidine (S3), after intracanal dressing (S4), and after 17% EDTA irrigation (S5). Cultivable bacteria recovered from the 5 stages were counted and identified by means of polymerase chain reaction assay (16S rRNA). Both groups had colony-forming unit counts significantly reduced after S2 (P < .05); however, no significant difference was found between the irrigants (S2 and S3, P = .99). No difference in bacteria counts was found between the intracanal medicaments used (P = .95). The most prevalent bacteria detected were Actinomyces naeslundii (66.67%), followed by Porphyromonas endodontalis, Parvimonas micra, and Fusobacterium nucleatum, which were detected in 33.34% of the root canals. An average of 2.13 species per canal was found, and no statistical correlation was observed between bacterial species and clinical/radiographic features. The microbial profile of infected immature teeth is similar to that of primarily infected permanent teeth. The greatest bacterial reduction was promoted by the irrigation solutions. The revascularization protocols that used the tested intracanal medicaments were efficient in reducing viable bacteria in necrotic immature teeth.

Insights into bacterial colony morphology evolution and diversification during infection development in cystic fibrosis

Tese de Doutoramento em Engenharia Biomédica.

Leukemic cluster growth in culture is an independent risk factor for acute myeloid leukemia and short survival in patients with myelodysplastic syndrome.

In patients with myelodysplastic syndrome (MDS) precursor cell cultures (colony-forming unit cells, CFU-C) can provide an insight into the growth potential of malignant myeloid cells. In a retrospective single-center study of 73 untreated MDS patients we assessed whether CFU-C growth patterns were of prognostic value in addition to established criteria. Abnormalities were classified as qualitative (i.e. leukemic cluster growth) or quantitative (i.e. strongly reduced/absent growth). Thirty-nine patients (53%) showed leukemic growth, 26 patients (36%) had strongly reduced/absent colony growth, and 12 patients showed both. In a univariate analysis the presence of leukemic growth was associated with strongly reduced survival (at 10 years 4 vs. 34%, p = 0.004), and a high incidence of transformation to AML (76 vs. 32%, p = 0.01). Multivariate analysis identified leukemic growth as a strong and independent predictor of early death (relative risk 2.12, p = 0.03) and transformation to AML (relative risk 2.63, p = 0.04). Quantitative abnormalities had no significant impact on the disease course. CFU-C assays have a significant predictive value in addition to established prognostic factors in MDS. Leukemic growth identifies a subpopulation of MDS patients with poor prognosis.

An HIV-1 clade C DNA prime, NYVAC boost vaccine regimen induces reliable, polyfunctional, and long-lasting T cell responses

The EuroVacc 02 phase I trial has evaluated the safety and immunogenicity of a prime-boost regimen comprising recombinant DNA and the poxvirus vector NYVAC, both expressing a common immunogen consisting of Env, Gag, Pol, and Nef polypeptide domain from human immunodeficiency virus (HIV)-1 clade C isolate, CN54. 40 volunteers were randomized to receive DNA C or nothing on day 0 and at week 4, followed by NYVAC C at weeks 20 and 24. The primary immunogenicity endpoints were measured at weeks 26 and 28 by the quantification of T cell responses using the interferon gamma enzyme-linked immunospot assay. Our results indicate that the DNA C plus NYVAC C vaccine regimen was highly immunogenic, as indicated by the detection of T cell responses in 90% of vaccinees and was superior to responses induced by NYVAC C alone (33% of responders). The vaccine-induced T cell responses were (a) vigorous in the case of the env response (mean 480 spot-forming units/10(6) mononuclear cells at weeks 26/28), (b) polyfunctional for both CD4 and CD8 T cell responses, (c) broad (the average number of epitopes was 4.2 per responder), and (d) durable (T cell responses were present in 70% of vaccinees at week 72). The vaccine-induced T cell responses were strongest and most frequently directed against Env (91% of vaccines), but smaller responses against Gag-Pol-Nef were also observed in 48% of vaccinees. These results support the development of the poxvirus platform in the HIV vaccine field and the further clinical development of the DNA C plus NYVAC C vaccine regimen

Fluorodeoxyuridine mediated cell cycle synchronization in S-phase increases the Auger radiation cell killing with 125I-iododeoxyuridine.

Aim: 125I-iododeoxyuridine is a potential Auger radiation therapy agent. Its incorporation in DNA of proliferating cells is enhanced by fluorodeoxyuridine. Here, we evaluated therapeutic activities of 125I-iododeoxyuridine in an optimized fluorodeoxyuridine pre-treatment inducing S-phase synchronization. Methods: After S-phase synchronization by fluorodeoxyuridine, cells were treated with 125I-iododeoxyuridine. Apoptosis analysis and S-phase synchronization were studied by flow cytometry. Cell survival was determined by colony-forming assay. Based on measured growth parameters, the number of decays per cell that induced killing was extrapolated. Results: Treatment experiments showed that 72 to 91% of synchronized cells were killed after 0.8 and 8 kBq/ml 125I-iododeoxyuridine incubation, respectively. In controls, only 8 to 38% of cells were killed by corresponding 125I-iododeoxyuridine activities alone and even increasing the activity to 80 kBq/ml gave only 42 % killing. Duplicated treatment cycles or repeated fluorodeoxyuridine pre-treatment allowed enhancing cell killing to >95 % at 8 kBq/ml 125I-iododeoxyuridine. About 50 and 160 decays per S-phase cells in controls and S-phase synchronization, respectively, were responsible for the observed cell killing at 0.8 kBq/ml radio-iododeoxyuridine. Conclusion: These data show the successful application of fluorodeoxyuridine that provided increased 125I-iododeoxyuridine Auger radiation cell killing efficacy through S-phase synchronization and high DNA incorporation of radio-iododeoxyuridine.

Cell-Cell interaction in erythropoiesis: role of human monocytes

Erythroid burst forming units (BFU-E) are proliferative cells present in peripheral blood and bone marrow which may be precursors of the erythroid colony forming cell found in the bone marrow. To examine the possible role of monocyte-macrophages in the modulation of erythropoiesis, the effect of monocytes on peripheral blood BFU-E proliferation in response to erythropoietin was investigated in the plasma clot culture system. Peripheral blood mononuclear cells from normal human donors were separated into four fractions. Fraction-I cells were obtained from the interface of Ficoll-Hypaque gradients (20-30% monocytes; 60-80% lymphocytes); fraction-II cells were fraction-I cells that were nonadherent to plastic (2-10% monocytes; 90-98% lymphocytes); fraction-III cells were obtained by incubation of fraction-II cells with carbonyl iron followed by Ficoll-Hypaque centrifugation (>99% lymphocytes); and fraction-IV cells represented the adherent population of fraction-II cells released from the plastic by lidocaine (>95% monocytes). When cells from these fractions were cultured in the presence of erythropoietin, the number of BFU-E-derived colonies was inversely proportional to the number of monocytes present (r = ¿0.96, P < 0.001). The suppressive effect of monocytes on BFU-E proliferation was confirmed by admixing autologous purified monocytes (fraction-IV cells) with fraction-III cells. Monocyte concentrations of ¿20% completely suppressed BFU-E activity. Reduction in the number of plated BFU-E by monocyte dilution could not account for these findings: a 15% reduction in the number of fraction-III cells plated resulted in only a 15% reduction in colony formation. These results indicate that monocyte-macrophages may play a significant role in the regulation of erythropoiesis and be involved in the pathogenesis of the hypoproliferative anemias associated with infection and certain neoplasia in which increased monocyte activity and monopoiesis also occur.

The development of a Defense System against Coxsackievirus B5 Infection in Suckling Mice

There are many viruses that are able to infect the alimentary tract of man. Little is known, however, about the mechanism of infection itself or the pathophysiology of the gut during infection. 'The research reported here is concerned with the differences in susceptibility among suckling mice of various ages inoculated by the intraperitoneal and intragastric routes. Since the normal mode of entry of many viruses to the gut is via the oral route, Coxsackievirus B5, a human enterovirus which does attack this way, was utilized. It is a non-tumor producing RNA virus that has been shown to act similarly in the mouse and human. The virus was pooled in HeLa cell cultures and titered by a plaquing assay in the same cell cultures. CD-l mice, 10, 14, 18, and 22 days old , were infected either orally or intraperitoneally with 5.0 x 10^10 (10 day old animals) and 1.0 x10^9 plaque forming units per animal. Dissections were done at 1 and 3 days post infection with samples of the blood, heart, liver, and gut being taken from each animal. Each sample was titered individually and the data presented as an average of six samples. As a result of previous work, it is known that the gut of a newborn mouse isn't able to decrease the concentration of the infecting dose and therefore provides no defense against an enteric infection with Coxsackievirus B5. In contrat, mature mice are able to reduce the amount of viral dissemination across the gut as well as inhibit replication after absorption has occurred. The results of this study indicate that there is a double barrier system developing in suckling mice that is involved with and directly related to the gastrointestinal tract The first part of this defense is the inhibition of penetration of virus across the gut when the primary site of' infection is the intestinal mucosa. This mechanism develops sometime around 20 to 22 days after birth. At about 16-18 days of age, suckling mice that were challenged intragastrically are able to stop active replication and initiate clearance of virus from the systemic circulation. There are many factors that might contribute to the marked decrease in susceptibility with age of suckling mice. Some of these or possibly a combination of these factors might explain the defense mechanisms described above, but to date, the chemistry or mechanical functioning of the gastrointestinal barrier to enteric viral infection is unknown.

Sensibilidade de Metarhizium anisopliae à temperatura e umidade em três tipos de solos

Este trabalho teve como objetivo investigar o efeito da temperatura e do teor de umidade do solo na sobrevivência de Metarhizium anisopliae (Metsch.) Sorok. em três tipos de solos. Foram utilizados o Latossolo Vermelho textura argilosa, Latossolo Vermelho textura média e Argissolo Vermelho Amarelo textura arenosa média. As temperaturas empregadas foram 21,5; 26,8 e 31,5°C, e os teores de umidade foram 35, 65 e 100% de saturação. A sobrevivência do fungo foi avaliada após zero, 20, 40, 60, 80, 100 e 120 dias de incubação em cada temperatura estudada. Na análise do efeito do teor de umidade, a sobrevivência foi avaliada após zero, 14, 28, 42, 56, 70, 84, 98 e 112 dias de incubação à temperatura de 27,0±1,0°C. em ambos os ensaios, foi determinado o número de unidades formadores de colônias (UFC) em placa de Petri. Houve influência significativa da temperatura e do teor de umidade na sobrevivência do fungo. O maior crescimento e a maior sobrevivência ocorreram nas temperaturas de 21,5 e 26,8°C, enquanto que, no solo incubado a 31,5°C, o fungo cresceu pouco, e a população declinou rapidamente. No teor de 65% de umidade, houve rápido crescimento do fungo, mas no 112° dia foi observado um declínio da população nos três tipos de solos. Nos teores de 35 e 100% de umidade, o crescimento foi menor, mas obteve-se maior sobrevivência do fungo no solo.

Antimycobacterial activity of some Brazilian indigenous medicinal drinks

Tuberculosis (TB) is a very serious problem worldwide and the increasing number of multiple drugs resistant TB cases makes the search for new anti-TB drugs an urgent need. Indigenous knowledge about the use of native plants to treat illnesses has contributed to the discovery of new medicines. In this study, the antimycobacterial activity of seven medicinal drinks was assessed: Ananas sativus (hydroalcoholic fruit extract), Aristolochia triangularis (aqueous and hydroalcoholic leaf, root and stem extracts), Bromelia antiacantha (hydroalcoholic fruit extract), Stryphnodendron adstringens (hydroalcoholic bark extract), Tabebuia ovellanedae (hydroalcoholic bark extract), Vernonia polyanthes (hydroalcoholic root extract), all used by the Vanuíre indigenous community in the treatment of respiratory diseases. The activity was evaluated by using a time-to-kill assay, in which Mycobacterium tuberculosis H37Rv was cultured on Löwenstein-Jensen medium, after thirty minutes, one, three, six, twelve and twenty-four hours contact of the bacteria with each drink. Within half to one hour contact, the hydroalcoholic drinks of A. triangularis, S. adstringens, T. ovellanedae and V. polyanthes reduced the bacterial growth by 2 orders of magnitude in CFU/mL, and all bacterial growth was absent after three hours contact. In contrast, no mycobactericidal effect was detected in the aqueous extract of A. triangularis or in the hydroalcoholic beverages of A. sativus and B. antiacantha, even after twenty-four hours contact.

Otimização da técnica de MIRU (Mycobacterial Interspersed Repetitive Units) para o estudo epidemiológico de pacientes com tuberculose

Pós-graduação em Biotecnologia - IQ

Serum bactericidal activity as indicator of innate immunity in pacu Piaractus mesopotamicus (Holmberg, 1887)

Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq)

Clinical Comparison of the Effectiveness of Single-file Reciprocating Systems and Rotary Systems for Removal of Endotoxins and Cultivable Bacteria from Primarily Infected Root Canals

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

Influência de diferentes métodos de esterilização de esmalte dental sobre a sua morfologia, composição química, estrutura e formação de biofilmes in vitro

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

Endodontic retreatment: clinical comparison of reciprocating systems versus rotary system in disinfecting root canals

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

The effect of blue light on periodontal biofilm growth in vitro

We have previously shown that blue light eliminates the black-pigmented oral bacteria Porphyromonas gingivalis, Prevotella intermedia, Prevotella nigrescens, and Prevotella melaninogenica. In the present study, the in vitro photosensitivity of the above black-pigmented microorganisms and four Fusobacteria species (Fusobacterium nucleatum ss. nucleatum, F. nucleatum ss. vincentii, F. nucleatum ss. polymorphum, Fusobacterium periodonticum) was investigated in pure cultures and human dental plaque suspensions. We also tested the hypothesis that phototargeting the above eight key periodontopathogens in plaque-derived biofilms in vitro would control growth within the dental biofilm environment. Cultures of the eight bacteria were exposed to blue light at 455 nm with power density of 80 mW/cm(2) and energy fluence of 4.8 J/cm(2). High-performance liquid chromatography (HPLC) analysis of bacteria was performed to demonstrate the presence and amounts of porphyrin molecules within microorganisms. Suspensions of human dental plaque bacteria were also exposed once to blue light at 455 nm with power density of 50 mW/cm(2) and energy fluence of 12 J/cm(2). Microbial biofilms developed from the same plaque were exposed to 455 nm blue light at 50 mW/cm(2) once daily for 4 min (12 J/cm(2)) over a period of 3 days (4 exposures) in order to investigate the cumulative action of phototherapy on the eight photosensitive pathogens as well as on biofilm growth. Bacterial growth was evaluated using the colony-forming unit (CFU) assay. The selective phototargeting of pathogens was studied using whole genomic probes in the checkerboard DNA-DNA format. In cultures, all eight species showed significant growth reduction (p < 0.05). HPLC demonstrated various porphyrin patterns and amounts of porphyrins in bacteria. Following phototherapy, the mean survival fractions were reduced by 28.5 and 48.2 % in plaque suspensions and biofilms, respectively, (p < 0.05). DNA probe analysis showed significant reduction in relative abundances of the eight bacteria as a group in plaque suspensions and biofilms. The cumulative blue light treatment suppressed biofilm growth in vitro. This may introduce a new avenue of prophylactic treatment for periodontal diseases.