Experimental infection of commercial layers using a Salmonella enterica sorovar Gallinarum strain: Blood serum components and histopathological changes

The purpose of the study was to evaluate the blood serum components and histopathological findings of commercial layers experimentally infected with Salmonella Gallinarum (SG), the microorganism responsible for the fowl typhoid. 180 commercial layers were distributed into three groups (G): G1 and G2 received 0.2mL of inoculum containing 3.3x10 8 and 3.3x10 5 CFU of resistant SG to the nalidix acid (Nal r)/mL, respectively, directly into their crops; G3 did not receive the inoculum (control group). The birds were inoculated when they were 5 days old and the euthanasia was performed 24 hours before and after infection and 3, 5, 7 and 10 days after the administration of the inoculum. In each day of collection, blood samples were obtained for biochemical tests of the blood serum besides macroscopic and histopathological examination of the birds. Data were submitted to analysis of variance by the SAS statistical program and the means were compared by Tukeýs test (P<0,05). In the serum biochemical profile it was observed that the infection interfered in the values of total protein, albumin, calcium, phosphorus, cholesterol, triglycerides, GGT and ALT in the infected groups. The macroscopic examination showed hepatomegaly, alteration of the hepatic color and hemorrhagic spots in the kidneys of animals from G1. The histopathology showed degeneration of hepatocytes in G1 and G2 although other lesions like multifocal hepatic necrosis and inflammatory infiltrate on the liver and kidneys were restricted to G1. The alterations were more evident on G1 which received a higher concentration of bacteria/mL when compared to G2. The results showed that the correlation between biochemical alterations and macroscopic and histopathological lesions can assist the comprehension of the pathophysiology of fowl typhoid, supplying important information for the diagnosis and prognosis of this disease.

Safety and efficacy of a Salmonella Gallinarum ΔcobSΔcbiA strain with potential to prevent chicken infections by Salmonella Gallinarum and Salmonella Enteritidis

New vaccine design techniques have allowed the development of effective vaccine strains against Salmonella infections in which the risks of reversion to the wild type and virulence is null. The mutant strain Salmonella Gallinarum ΔcobSΔcbiA was previously shown to be avirulent in chickens. In this study, this strain was tested as a vaccine against Salmonella Gallinarum (SG) and S. Enteritidis (SE) infections, and its protection levels, safety and possible risks of reversion to virulence after vaccination of layers were evaluated. Birds were vaccinated at five days of age or at five and 25 days of age. At 45 days of age, brown and white layers were challenged with SG and SE wild strains, respectively. Two assays to test the possibility of reversion to virulence were performed. Five successive bacterial passages in brown layers were carried out in the first assay. In the second assay, brown layers received a ten-fold concentrated inoculum of the SGΔcobSΔcbiA strain and were evaluated for clinical signs and mortality. In both experiments, no birds that received the inoculation of the attenuated strain died. Additionally, the use of the mutant strain as a vaccine provided good protection levels against both challenge strains.

Densidade de Enterococcus sp em águas recreacionais e areias de praias do município de São Vicente-SP, Brasil e sua relação com parâmetros abióticos

Coastal cities attract a large number of tourists for their beautiful landscape and recreational activities, increasing the municipalities' income source. Thus, a need is clear for the establishment of beach quality monitoring programs to ensure bathers health. Although there is an effective monitoring program for recreational waters in Brazil there are no programs to certify the quality of beach sands. In this sense, the aim of this work was to analyze the density of bacteria from the genus Enterococcus in both sand and water from two beaches from São Vicente, São Paulo (Brazil) and correlate these densities to abiotic parameters such as: temperature, salinity, particle size, organic matter and tides). Water and sand samples were collected during February 2006 on the beaches of Gonzaguinha e Ilha Porchat and bacterial densities were determined by membrane filter technique. Temperature and salinity were measured in situ with a thermometer and a refractometer while particle size and organic matter were determined according to methods described by Suguio and Dean. There were significant differences between densities found in water and sand (p=0.004), being approximately 20 times higher in Gonzaguinha's beach sands. Similar results were found for Ilha Porchat beach samples, being the densities found in sand 3 times greater than those found in water. Both beaches showed a negative correlation between bacterial densities and salinity and temperature, suggesting a deleterious effect of these parameters on the bacterial community. On the other hand, no correlation was found between organic matter and particle size and bacterial densities. This work demonstrates that there is a great necessity for monitoring beach sand since the evaluation of beach quality is an important part of integrated coastal management programs.

Photodynamic and Antibiotic Therapy Impair the Pathogenesis of Enterococcus faecium in a Whole Animal Insect Model

Enterococcus faecium has emerged as one of the most important pathogens in healthcare-associated infections worldwide due to its intrinsic and acquired resistance to many antibiotics, including vancomycin. Antimicrobial photodynamic therapy (aPDT) is an alternative therapeutic platform that is currently under investigation for the control and treatment of infections. PDT is based on the use of photoactive dye molecules, widely known as photosensitizer (PS). PS, upon irradiation with visible light, produces reactive oxygen species that can destroy lipids and proteins causing cell death. We employed Galleria mellonella (the greater wax moth) caterpillar fatally infected with E. faecium to develop an invertebrate host model system that can be used to study the antimicrobial PDT (alone or combined with antibiotics). In the establishment of infection by E. faecium in G. mellonella, we found that the G. mellonella death rate was dependent on the number of bacterial cells injected into the insect hemocoel and all E. faecium strains tested were capable of infecting and killing G. mellonella. Antibiotic treatment with ampicillin, gentamicin or the combination of ampicillin and gentamicin prolonged caterpillar survival infected by E. faecium (P = 0.0003, P = 0.0001 and P = 0.0001, respectively). In the study of antimicrobial PDT, we verified that methylene blue (MB) injected into the insect followed by whole body illumination prolonged the caterpillar survival (P = 0.0192). Interestingly, combination therapy of larvae infected with vancomycin-resistant E. faecium, with antimicrobial PDT followed by vancomycin, significantly prolonged the survival of the caterpillars when compared to either antimicrobial PDT (P = 0.0095) or vancomycin treatment alone (P = 0.0025), suggesting that the aPDT made the vancomycin resistant E. faecium strain more susceptible to vancomycin action. In summary, G. mellonella provides an invertebrate model host to study the antimicrobial PDT and to explore combinatorial aPDT-based treatments.

Comparative analysis of Enterococcus faecalis biofilm formation on different substrates

Introduction: The aim of this study was to compare Enterococcus faecalis biofilm formation on different substrates. Methods: Cell culture plates containing growth medium and E. faecalis (ATCC 29212) were used to grow biofilm on bovine dentin, gutta-percha, hydroxyapatite, or bovine bone. Substrates were incubated at 37°C for 14 or 21 days, and the medium was changed every 48 hours. After the growth induction periods, specimens (n = 5 per group and per induction period) were stained by using Live/Dead, and the images were analyzed under a confocal microscope. The total biovolume (μm3), live bacteria biovolume (μm3), and substrate coverage (%) were quantified by using the BioImage-L software. Results obtained were analyzed by nonparametric tests (P =.05). Results: Biofilm formation was observed in all groups. Gutta-percha had the lowest total biovolume at 14 days (P <.05) and hydroxyapatite the highest at 21 days (P <.05). No significant difference was observed in green biovolume at 14 days. At 21 days, however, hydroxyapatite had the highest volume (P <.05). The percentages of coverage were similar among all substrates at 21 days (P >.05), but at 14 days, bovine bone presented the highest coverage (P <.05). Conclusions: E. faecalis was capable of forming biofilm on all substrates during both growth periods; hydroxyapatite presented the highest rates of biofilm formation. The type of substrate influenced the biofilm characteristics, according to the parameters evaluated. © 2013 American Association of Endodontists.

Ex-vivo Effect of Intracanal Medications Based on Ozone and Calcium Hydroxide in Root Canals Contaminated with Enterococcus faecalis

This ex vivo study evaluated the antibacterial effect of intracanal medications in root canals contaminated with Enterococcus faecalis. Fifty single-rooted human teeth were contaminated with E. faecalis (ATCC 29212) and incubated at 37°C for 21 days. The specimens were randomly divided into 5 groups according to the intracanal medication used: OZ-PG: ozonized propylene glycol; CH/CPMC: calcium hydroxide/camphorated paramonochlorophenol; OZ-PG/CH ozonized PG/CH; PC: positive control group (no medication); and NC: negative control group (no contamination). The samples were collected after 7 days (post-medication) and 14 days (final). Bacterial growth was checked by counting the colony-forming units (CFU). OZ-PG and CH/CPMC reduced significantly the CFU counts compared with PC in the post-medication and final samples, with no statistically significant differences among them. On the other hand, OZ-PG/CH did not reduce significantly the number of bacteria compared with PC. In conclusion, among the evaluated medications OZ-PG and CH/CPMC were the most effective against E. faecalis.

In vitro antimicrobial activity of auxiliary chemical substances and natural extracts on Candida albicans and Enterococcus faecalis in root canals

Objective: The aim of this study was to evaluate the antimicrobial activity of auxiliary chemical substances and natural extracts on Candida albicans and Enterococcus faecali's inoculated in root canals. Material and Methods: Seventy-two human tooth roots were contaminated with C. albicans and E. faecalis for 21 days. The groups were divided according to the auxiliary chemical substance into: G1) 2.5% sodium hypochlorite (NaOCl), G2) 2% chlorhexidine gel (CHX), G3) castor oil, G4) glycolic Aloe vera extract, g5) glycolic ginger extract, and G6) sterile saline (control). The samples of the root canal were collected at different intervals: confirmation collection, at 21 days after contamination; 1st collection, after instrumentation; and 2nd collection, seven days after instrumentation. Microbiological samples were grown in culture medium and incubated at 37°C for 48 hours. Results: The results were submitted to the Kruskal-Wallis and Dunn (5%) statistical tests. NaOCl and CHX completely eliminated the microorganisms of the root canals. Castor oil and ginger significantly reduced the number of CFU of the tested bacteria. Reduction of CFU/mL at the 1st and 2nd collections for groups G1, G2, G3 and G4 was greater in comparison to groups G5 and G6. Conclusion: It was concluded that 2.5% sodium hypochlorite and 2% chlorhexidine gel were more effective in eliminating C. albicans and E. faecalis, followed by the castor oil and glycolic ginger extract. The Aloe vera extract showed no antimicrobial activity.

Polymerase chain reaction assay based on ratA gene allows differentiation between Salmonella enterica subsp. enterica serovar Gallinarum biovars Gallinarum and Pullorum

Salmonella Pullorum and Salmonella Gallinarum are classified as biovars of Salmonella enterica subsp. enterica serovar Gallinarum. These salmonellae are the causative agents of Pullorum disease and fowl typhoid, respectively, and are widely distributed throughout the world. Although many developed countries have eradicated these diseases from commercial poultry, they are still the cause of significant economic loss in developing countries. When serovar Gallinarum is isolated, it is difficult to immediately differentiate between biovars because they are antigenically identical by serotyping. However, they cause distinct diseases with different epidemiology, and therefore it is important to differentiate them. This may be done biochemically but takes 2 to 3 days. In the present study, S. Pullorum and S. Gallinarum whole genomes were compared, and 1 genomic region of difference, which is part of the ratA gene, was chosen as a molecular marker for a polymerase chain reaction assay to differentiate rapidly between these organisms. In all, 26 strains of S. Gallinarum and 17 S. Pullorum strains were tested and successfully differentiated by the assay. © 2013 The Author(s).

Experimental infection of commercial layers with wild or attenuated Salmonella Gallinarum mutant strains: Anatomic pathology, total blood cell count and serum protein levels

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

Estudo comparativo in vitro da suscetibilidade à ciprofloxacina e à vancomicina em linhagens de Enterococcus faecalis isoladas de processos infecciosos

Pós-graduação em Doenças Tropicais - FMB

Prevalência e suscetibilidade aos antibióticos de enterococcus spp. isoladas da cavidade bucal humana

Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES)

Associação de enterococcus faecalis, candida albicans e escherichia coli em canais radiculares e avaliação dos efeitos de extratos naturais sobre os microrganismos e endotoxinas

Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES)

O emprego da metodologia de superfície de resposta no desenvolvimento de um novo produto simbiótico, fermentado com Enterococcus faecium CRL 183 e Lactobacillus helveticus ssp jugurti 416, à base de extratos aquosos de soja e de Yacon (Smallanthus sonchifolius)

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

Avaliação do simbiótico fermentado com Enterococcus faecium CRL 183 e Lactobacillus helveticus ssp jugurti 416, à base de extratos aquosos de soja e de yacon (Smallanthus sonchifolius) no controle do desenvolvimento do Diabetes Mellitus

Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES)

Ação in vitro do extrato glicólico de gengibre e medicamentos sobre Candida albicans, Enterococcus faecalis, Escherichia coli e sua endotoxina em canais radiculares

Pós-graduação em Odontologia Restauradora - ICT