Escherichia coli de adesão difusa (DAEC) isoladas de crianças e de adultos constituem duas populações diferentes

Tese (doutorado)—Universidade de Brasília, Instituto de Biologia, Departamento de Biologia Celular, Programa de Pós-Graduação em Biologia Molecular, 2013.Escherichia coli de adesão difusa (DAEC) é considerada uma categoria diarreiogênica de E. coli. Essas cepas são identificadas pelo padrão de adesão em cultura de células. A maioria das cepas DAEC possui adesinas da família Afa/Dr. Sua patogenia tem sido muito discutida, embora alguns fatores de virulência já tenham sido propostos. O objetivo principal deste trabalho foi comparar cepas DAEC Afa/Dr isoladas de crianças e de adultos, a partir da análise de características relacionadas à virulência e à formação de biofilmes. Cepas DAEC Afa/Dr oriundas de crianças e adultos apresentaram marcantes diferenças na distribuição de características. Na coleção de adultos, uma nova adesina da família Afa/Dr foi associada aos casos de diarreia. Cepas de crianças, principalmente as de controles, apresentaram grande diversidade de adesinas Afa/Dr, nenhuma delas associada à diarreia. As cepas DAEC mostraram habilidade aumentada de formação de biofilme quando associadas a uma cepa de Citrobacter freundii. O aumento da formação de biofilme foi associado a maior capacidade de adesão a células cultivadas, indicando que cepas DAEC podem apresentar maior capacidade de colonização, dependendo dos parceiros, o que por sua vez, poderia aumentar a efetividade de uma toxina, como SAT - encontrada em número significativamente superior em cepas isoladas de casos de diarreia em crianças. A expressão de curli em meio CRI por cepas DAEC produziu um fenótipo adicional aos conhecidos em Enterobacteriacea. A frequência de curli foi significativamente associada aos casos de diarreia em adultos, sugerindo que curli é um possível fator de virulência nessas bactérias. Genes do TTSS e sorogrupos clássicos de EPEC foram encontrados apenas nas cepas de crianças. Somados, os resultados indicam que cepas DAEC Afa/Dr isoladas de adultos e de crianças constituem duas populações diferentes. _____________________________________________________________________________________________________ ABSTRACTDiffusely adherent Escherichia coli (DAEC) is considered a diarrheagenic category of E. coli. Those strains are identified by their adhesion pattern in cell cultures. Most DAEC strains possess adhesins belonging to the Afa/Dr family. Their pathogenicity has been widely discussed, although some virulence factors have already been proposed. This work’s main goal is to compare DAEC Afa/Dr strains isolated from children and adults, by analyzing characteristics related to virulence and biofilm formation. DAEC Afa/Dr strains from children and adults present remarkable differences in the distribution of the studied characteristics. Among strains from adults, a new adhesin from the Afa/Dr family was associated to cases of diarrhea. Strains from children, especially those isolated from control, present great diversity of Afa/Dr adhesins, none associated with diarrhea. DAEC strains showed an increase in biofilm formation when associated to a strain of Citrobacter freundii. The increase in biofilm formation was associated to a greater adhesion capacity in cultivated cells, indicating that DAEC strains can present a greater capacity for colonization depending on its partners, which, in turn, could increase the effectiveness of toxins, such as SAT - found in significantly higher numbers in strains isolated from cases of diarrhea in children. The expression of curli in the CRI medium produced an additional phenotype, as well as others previously described for Enterobacteriacea. The prevalence of curli was significantly associated to cases of diarrhea in adults, indicating that curli could be a possible virulence factor in those bacteria. Genes from the TTSS and classical EPEC serogroups were found exclusively in strains from children. Taken together, these results indicate that DAEC Afa/Dr strains isolated from adults and children constitute two different populations

Diffusely adherent Escherichia coli strains isolated from children and adults constitute two different populations

Background: Diffusely adherent Escherichia coli (DAEC) have been considered a diarrheagenic category of E. coli for which several potential virulence factors have been described in the last few years. Despite this, epidemiological studies involving DAEC have shown inconsistent results. In this work, two different collections of DAEC possessing Afa/Dr genes, from children and adults, were studied regarding characteristics potentially associated to virulence. Results: DAEC strains were recovered in similar frequencies from diarrheic and asymptomatic children, and more frequently from adults with diarrhea (P < 0.01) than from asymptomatic adults. Association with diarrhea (P < 0.05) was found for SAT-positive strains recovered from children and for curli-positive strains recovered from adults. Mixed biofilms involving DAEC and a Citrobacter freundii strain have shown an improved ability to form biofilms in relation to the monocultures. Control strains have shown a greater diversity of Afa/Dr adhesins and higher frequencies of cellulose, TTSS, biofilm formation and induction of IL-8 secretion than strains from cases of diarrhea in children. Conclusions: DAEC strains possessing Afa/Dr genes isolated from children and adults represent two different bacterial populations. DAEC strains carrying genes associated to virulence can be found as part of the normal microbiota present in asymptomatic children

hsCRP and E-Selectin as Markers of Endothelial Dysfunction in Children with Type 1 Diabetes Mellitus

Introduction: This study investigated the levels of inflammatory biomarkers in healthy children and those diagnosed with type 1 diabetes (T1DM), some of whom were affected by endothelial dysfunction (ED), characterized by increased inflammation and reduced vasodilatation. Methods: Thirty-one T1DM children showing no symptoms of vascular diseases and diagnosed by ultrasound techniques as ED-positive (T1DM-ED) or negative (T1DM), and 58 sex-age-matched healthy children were investigated for circulating levels of E-selectin, s-ICAM and s-VCAM, MMP-9, high-sensitivity C-reactive protein (hsCRP), and IL-6. Results: No differences were observed in s-ICAM, MMP-9, and IL-6 levels between case and control groups. Significantly higher levels of s-VCAM (p= 0.0001) were found in the T1DM (1359.1 ± 273 ng/mL) and T1DM-ED (1358.2 ± 112 ng/mL) groups; (control - 828.5 ± 212 ng/mL). Higher levels of E-selectin (p = 0.001) were found in the T1DM-ED group (331.2 ± 77 ng/mL); (control - 222.2 ± 74 ng/mL). The values of hsCRP were higher (p = 0.002) in the T1DM-ED group (0.36 ± 0.2 mg/L) relative to control (0.15 ± 0.1 mg/L) and T1DM (0.19 ± 0.2 mg/L). The results suggest that E-selectin and hsCRP can be useful markers of ED in children with T1DM.publishersversionpublishe

Inhibitory effects of Lactobacillus casei Shirota against both Candida auris and Candida spp. isolates that cause vulvovaginal candidiasis and are resistant to antifungals

Background Vulvovaginal candidiasis (VVC), the second leading cause of genital infection in women of reproductive age, is caused by yeasts of the genus Candida. Treatment is usually empirical and performed with azoles, which have shown increasing ineffectiveness due to resistance from these species. This therapeutic challenge has led to the search for new treatment strategies. Lactobacillus spp. produce several components with microbicidal effects, such as lactic acid. These species are the main components of a healthy vaginal microbiota and have been used as probiotics. The aim of this work was to investigate the in vitro inhibitory effects of Lactobacillus casei Shirota on both the Candida spp. that cause VVC and on C. auris. Methods The microbicidal effects of L. casei Shirota on the main VVC-causing species, C. albicans, C. tropicalis, C. norvegensis and C. parapsilosis, in addition to C. auris were investigated by counting the Colony-forming Units (CFUs) after cocultivation. The antifungal activity of lactic acid against these Candida strains was assessed using the microtiter broth dilution method to determine the minimum inhibitory concentrations (MICs). The effects of L. casei Shirota on hyphal and early biofilm formation was measured by optical microscopy. Results L. casei Shirota showed inhibitory action against all tested Candida spp., ranging from 66.9 to 95.6% inhibition depending on the species. This inhibition is possibly related to the production of lactic acid, since lactic acid has shown microbicidal action against these same Candida spp. at a concentration of 5 mg/mL, which corresponds to half of the normal physiological concentration. In addition, L. casei Shirota was able to reduce the formation of C. albicans hyphae and early biofilms, showing strong anti-Candida effects. Conclusions These results suggest that L. casei Shirota has antifungal activity against the Candida species that cause VVC. L. casei also has microbicidal action against C. auris

Oral or topical administration of L-arginine changes the expression of TGF and iNOS and results in early wounds healing

PURPOSE: To evaluate the contribution of L-arginine oral or topical rout of administration in the surgical wound healing process. METHODS: L-arginine was orally or topically administrated to mice after a laparotomy model procedure. The wounds were analyzed to evaluate the granulation tissue by HE analysis, collagen deposition, iNOS and cytokines production by immunochemisyry on wound progress. Mice used in this model were healthy, immunosupressed or diabetic and all of them were treated with different concentration of L-arginine and rout of administration. RESULTS: Suggested that groups treated with L-arginine orally or topically improved wound repair when compared with non-treatad mice. L- arginine treatment stimulated TGF-β and restricted NO production leading to a mild Th1 response and collagen deposition in injured area, when it was orally administrated. Topical administration decreased IL-8 and CCR1 expression by wound cells but did not interfere with TNF-α and IL-10 production, ratifying the decrease of inflammatory response, the oral administration however, presented a higher iNOS and TGF-β expression then. L-arginine treatment also improved the improved the wound healing in immunosupressed or diabetic mice. CONCLUSION: L-arginine administrated orally or topically can be considered an important factor in the recuperation of tissues