Tripanosomíase americana (doença de Chagas) em ratos e camundongos convencionais e isentos de germes

Germfree (GF) and conventional (CV) CFW (LOB) mice and Wistar and Sprague-Dawley rats were infected with Trypanosoma cruzi. The disease was more severe in the GF than in the CV animals as revealed by: (1) an earlier and more intense parasitemia; (2) a more precocious mortality; (3) a twice enlarged spleen: (4) a more intense cell and tissue parasitism; (5) visceral signs of cardiac failure.Camundongos CFW (LOB) e ratos Wistar e Sprague-Dawley isentos de germes (GF) e convencionais (CV) foram infectados com Trypanosoma cruzi. A doença foi mais grave nos animais GF do que nos CV, o que foi demonstrado por: (1) uma parasitemia mais precoce e mais intensa: (2) uma mortalidade mais precoce: (3) baço duas vezes maior; (4) um parasitismo celular e tissular mais intenso; (5) sinais viscerais de insuficiência cardíaca

Genetic transformation of novel isolates of chicken Lactobacillus bearing probiotic features for expression of heterologous proteins: a tool to develop live oral vaccines

BACKGROUND: The use of lactic acid bacteria as vehicles to delivery antigens to immunize animals is a promising issue. When genetically modified, these bacteria can induce a specific local and systemic immune response against selected pathogens. Gastric acid and bile salts tolerance, production of antagonistic substances against pathogenic microorganisms, and adhesive ability to gut epithelium are other important characteristics that make these bacteria useful for oral immunization. RESULTS: Bacteria isolated on de Man, Rogosa and Sharpe medium (MRS) from different gastrointestinal portions of broiler chicks were evaluated for their resistance to artificial gastric acid and bile salts, production of hydrogen peroxide, and cell surface hydrophobicity. Thirty-eight isolates were first typed at species level by PCR amplification of 16S-23S rRNA intergenic spacers using universal primers that anneal within 16S and 23S genes, followed by restriction digestion analyses of PCR amplicons (PCR-ARDRA). An expression cassette was assembled onto the pCR2.1-Topo vector by cloning the promoter, leader peptide, cell wall anchor and terminator sequences derived from the laminin binding S-layer protein gene of L. crispatus strain F5.7 (lbs gene). A sequence encoding the green fluorescent protein (GFP) was inserted as reporter gene, and an erythromycin resistance gene was added as selective marker. All constructs were able to express GFP in the cloning host E. coli XL1-Blue and different Lactobacillus strains as verified by FACS and laser scanning confocal microscopy. CONCLUSION: Lactobacillus isolated from gastrointestinal tract of broiler chickens and selected for probiotic characteristics can be genetically modified by introducing an expression cassette into the lbs locus. The transformed bacteria expressed on its cell wall surface different fluorescent proteins used as reporters of promoter function. It is possible then that similar bacterial model expressing pathogen antigens can be used as live oral vaccines to immunize broilers against infectious diseases

Interaction of Saccharomyces boulardii with Salmonella enterica Serovar Typhimurium Protects Mice and Modifies T84 Cell Response to the Infection

BACKGROUND: Salmonella pathogenesis engages host cells in two-way biochemical interactions: phagocytosis of bacteria by recruitment of cellular small GTP-binding proteins induced by the bacteria, and by triggering a pro-inflammatory response through activation of MAPKs and nuclear translocation of NF-kappaB. Worldwide interest in the use of functional foods containing probiotic bacteria for health promotion and disease prevention has increased significantly. Saccharomyces boulardii is a non-pathogenic yeast used as a probiotic in infectious diarrhea. METHODOLOGY/PRINCIPAL FINDINGS: In this study, we reported that S. boulardii (Sb) protected mice from Salmonella enterica serovar Typhimurium (ST)-induced death and prevented bacterial translocation to the liver. At a molecular level, using T84 human colorectal cancer cells, we demonstrate that incubation with Sb before infection totally abolished Salmonella invasion. This correlates with a decrease of activation of Rac1. Sb preserved T84 barrier function and decreased ST-induced IL-8 synthesis. This anti-inflammatory effect was correlated with an inhibitory effect of Sb on ST-induced activation of the MAPKs ERK1/2, p38 and JNK as well as on activation of NF-kappaB. Electron and confocal microscopy experiments showed an adhesion of bacteria to yeast cells, which could represent one of the mechanisms by which Sb exerts its protective effects. CONCLUSIONS: Sb shows modulating effects on permeability, inflammation, and signal transduction pathway in T84 cells infected by ST and an in vivo protective effect against ST infection. The present results also demonstrate that Sb modifies invasive properties of Salmonella

Strongyloides venezuelensis: efeito de antimicrobiano e imunossupressor no curso da infecção em camundongos da linhagem AKR/J

Groups of AKR/J strain of mice infected by Strongyloides venezuelensis and treated with Ceftazidima, Dexametasona or with both drugs concomitantly had been killed on 3rd, 7th and 12th day after infection and their lungs and intestines were processed for histological studies. In lungs of all infected groups, it was verified an inflammatory infiltrated (neutrophils and mononuclear cells) in the third day after the infection. In 7th and 12th day after the infection, the inflammatory reaction becomes reduced in control and antimicrobial treated groups, but not in immunosuppressed animals (with or without antimicrobial treatment). Analysis of the duodenal mucosa confirmed the presence of parasites in all groups of animals. On the 7th day after infection it was observed significant alterations in the small intestine of control (infected) and infected and treated with antimicrobial groups with presence of inflammatory foci, constituted by mononuclear and eosinophils in mucosa, associate to a large amount of parasites, mainly in the region of the epithelium and sub epithelium. Two others groups (infected and immunosuppressed mice with or without antimicrobial), did not present inflammatory process. Goblet cells were less evident in mucosa suggesting a reduction in mucous production. In the 12th day of the infection, the treated with antimicrobial and control groups presented a reduced number of parasites and a discrete inflammatory reaction in the small intestine while the immunosuppressed groups showed more parasites in duodenum. The permanence of the infection by S. venezuelensis in immunosuppressed animals was prolonged in relation to other groups, remaining until the 49th day after infection. The results indicate that the interference of treatments in the population of intestinal microbiota favours the parasitism by Strongyloides venezuelensis.Camundongos da linhagem AKR/J infectados pelo Strongyloides venezuelensis e tratados com Ceftazidima, Dexametasona ou com ambas as drogas foram sacrificados no terceiro, sétimo e décimo segundo dia após infecção. Pulmões e intestino delgado foram processados para histologia. Verificaram-se, nos pulmões dos quatro grupos infectados, infiltrados inflamatórios (neutrófilos e mononucleares) no terceiro dia após a infecção. No sétimo e décimo segundo dia após a infecção, o processo inflamatório se torna reduzido nos animais controle e tratados com antimicrobiano, mas não nos animais imunossuprimidos (com ou sem tratamento com antimicrobiano). A análise da mucosa duodenal confirmou a presença de parasitos em todos os grupos de animais. No 7º dia após a infecção, foram observadas alterações significativas no duodeno dos grupos controle (infectados) e dos infectados e tratados com antimicrobiano, com presença de infiltrado inflamatório, constituído de mononucleares e eosinófilos na mucosa, associada a uma maior quantidade de parasitos, principalmente na região do epitélio e sub-epitélio. Os outros dois grupos (camundongos infectados e imunossuprimidos com ou sem antimicrobiano), não apresentaram processo inflamatório. A mucosa apresentou células caliciformes menos evidentes, sugerindo uma redução de produção de muco. No décimo segundo dia da infecção, os grupos não tratados e tratados apenas com antimicrobiano apresentaram um número reduzido de parasitos e um discreto processo inflamatório no duodeno, enquanto os grupos tratados com imunossupressor (com ou sem antimicrobiano) mostraram presença de um maior número do parasito no duodeno. A permanência da infecção do S. venezuelensis foi mais prolongada nos animais imunossuprimidos em relação aos demais grupos, permanecendo até o 490dia após infecção. Os resultados indicam que a interferência dos tratamentos no equilíbrio da microbiota intestinal favoreceu o parasitismo pelo S. venezuelensis

Oral treatment with Saccharomyces cerevisiae strain UFMG 905 modulates immune responses and interferes with signal pathways involved in the activation of inflammation in a murine model of typhoid fever

AbstractSalmonella spp. are Gram-negative, facultative, intracellular pathogens that cause several diarrheal diseases ranging from self-limiting gastroenteritis to typhoid fever. Previous results from our laboratory showed that Saccharomyces cerevisiae strain UFMG 905 isolated from ‘cachaça’ production presented probiotic properties due to its ability to protect against experimental infection with Salmonella enterica serovar Typhimurium. In this study, the effects of oral treatment with S. cerevisiae 905 were evaluated at the immunological level in a murine model of typhoid fever. Treatment with S. cerevisiae 905 inhibited weight loss and increased survival rate after Salmonella challenge. Immunological data demonstrated that S. cerevisiae 905 decreased levels of proinflammatory cytokines and modulated the activation of mitogen-activated protein kinases (p38 and JNK, but not ERK1/2), NF-κB and AP-1, signaling pathways which are involved in the transcriptional activation of proinflammatory mediators. Experiments in germ-free mice revealed that probiotic effects were due, at least in part, to the binding of Salmonella to the yeast. In conclusion, S. cerevisiae 905 acts as a potential new biotherapy against S. Typhimurium infection due to its ability to bind bacteria and modulate signaling pathways involved in the activation of inflammation in a murine model of typhoid fever

Variability of cutaneous and nasal population levels between patients colonized and infected by multidrug-resistant bacteria in two Brazilian intensive care units

Objective: To compare cutaneous and nasal population levels between patients colonized and infected by multidrug-resistant organisms in two intensive care units. Methods: A prospective cohort study was performed in adult intensive care units of two hospitals in Belo Horizonte, Brazil (April 2012 to February 2013). Clinical and demographic data were first collected by reviewing patients’ charts. Then, samples collected with nasal, groin, and perineum swabs were cultivated in selective media for 48 h at 37°C. After isolation, determination of antimicrobial susceptibility and biochemical identification were performed. Results: A total of 53 cases of colonization were observed by the following bacteria in decreasing frequencies: imipenem-resistant Acinetobacter baumannii (50.9%), vancomycin-resistant Enterococcus faecalis (43.4%), extended-spectrum beta-lactamase–producing Klebsiella pneumoniae (37.7%), imipenem-resistant Pseudomonas aeruginosa (32.1%), oxacillin-resistant Staphylococcus aureus (7.5%), and imipenem-resistant Klebsiella pneumoniae (5.7%). Among these colonization cases, 26 (49.0%) were followed by infection with bacteria phenotypically similar to those of the colonization. A relation between high population levels of colonization by most of the multidrug-resistant organisms at anatomical sites and a subsequent infection was observed. After colonization/infection, bacterial population levels decreased progressively and spontaneously until disappearance by day 45 in all the anatomical sites and for all the multidrug-resistant organisms. Conclusion: There was a correlation between high population levels of colonization by multidrug-resistant organisms at anatomical sites and a subsequent infection. Reduction in multidrug-resistant organism populations after colonization at anatomical sites could be a preventive measure to reduce evolution to infection as well as transmission of these bacteria between patients in intensive care unit

Protection by Lactobacillus acidophilus UFV-H2B20 against experimental oral infection with Salmonella enterica subsp. enterica Ser. Typhimurium in gnotobiotic and conventional mice Proteção por Lactobacillus acidophilus UFV-H2B20 contra o desafio oral experimental com Salmonella enterica subsp. enterica Ser. Typhimurium em camundongos gnotobióticos e convencionais

The ability of Lactobacillus acidophilus UFV-H2B20 to antagonize Salmonella enterica subsp. enterica ser. Typhimurium and to reduce the pathological consequences for the host was determining using conventional and gnotobiotic animals. Conventional NIH mice received daily by gavage a 0.1 ml suspension containing about 10(8) cfu L. acidophilus UFV-H2B20 and germfree animals received a single 0.1 ml dose. The gnotobiotic and conventional groups were infected orally with 10² and 10(5) cfu of S. Typhimurium, respectively, 7 days after the beginning of treatment. Control groups were treated with sterile saline instead of Lactobacillus. Survival data showed a protective effect against the pathogenic bacteria in both conventional and gnotobiotic Lactobacillus-treated mice. L. acidophilus UFV-H2B20 colonized the digestive tract of gnotobiotic mice and the number of viable cells ranged from 10(9) to 10(10) cfu/g of faeces. In both experimental and control gnotobiotic animals, S. Typhimurium became rapidly established at a level ranging from 10(8) to 10(10) cfu/g of faeces and remained at high levels until the animals died or were sacrificed. In conclusion, the previous treatment of mice with L. acidophilus UFV-H2B20 protects the animals against the experimental infection with S. Typhimurium but this protection was not due to the reduction of the pathogenic populations in the intestines.<br>A capacidade de Lactobacillus acidophilus UFV-H2B20 de antagonizar Salmonella enterica subsp. enterica ser. Typhimurium, e de reduzir as conseqüências patológicas para o hospedeiro foram determinadas em animais convencionais e gnotobióticos. Camundongos NIH convencionais receberam diariamente, por via oral, 0,1 ml de uma suspensão contendo em torno de 10(8) ufc de L. acidophilus UFV-H2B20 e os animais sem germes receberam uma única dose de 0,1 ml. Os grupos gnotobióticos e convencionais foram desafiados oralmente com, respectivamente, 10² e 10(5) ufc de S. Typhimurium 7 dias após o início do tratamento. Os grupos controles foram tratados com salina estéril em vez do Lactobacillus. Dados de sobrevida mostraram um efeito protetor contra a bactéria patogênica em ambos os grupos convencional e gnotobiótico tratados com o Lactobacillus. L. acidophilus UFV-H2B20 colonizou o trato digestivo dos camundongos gnotobióticos e o número de células víaveis flutuou entre 10(9) e 10(10) ufc/g de fezes. Em ambos os grupos experimental e controle, S. Typhimurium se estabeleceu rapidamente numa faixa de 10(8) a 10(10) ufc/g de fezes e se manteve em níveis elevados até os animais morreram ou serem sacrificados. Em conclusão, o tratamento prévio de camundongos com L. acidophilus UFV-H2B20 protege os animais contra a infecção experimental com S. Typhimurium mas essa proteção não se deve à uma redução das populações patogênicas nos intestinos

Effect of the trehalose levels on the screening of yeast as probiotic by in vivo and in vitro assays Efeito da trealose na seleção de leveduras para uso como probióticos utilizando testes in vitro e in vivo

Probiotics are viable defined microorganisms (bacteria or yeasts) that exert a beneficial effect on the health of the host when ingested in adequate amounts. Screening for such biotherapeutic agents is commonly performed by in vitro assays simulating gastrointestinal environment to determine the ability to survive in the digestive tract. In the present study, the possibility of extrapolation of data obtained in in vitro assays to in vivo conditions was studied using five Saccharomyces cerevisiae strains isolated from Brazilian Atlantic rain forest. Trehalose contents and survival after exposure to a combination of physiological stresses generally found in the gastrointestinal tract of humans were determined for the five yeasts and compared to the behavior of Saccharomyces boulardii, a well-known probiotic. The results were completed with the colonization capacity of the gastrointestinal tract of gnotobiotic mice by these yeast strains. Some results obtained by in vitro assays are not confirmed by in vivo experiments, indicating that the extrapolation cannot be always done.<br>Probióticos são definidos como microrganismos (bactérias e leveduras) que exercem um efeito benéfico na saúde do hospedeiro quando ingeridos em quantidades adequadas. A seleção desses agentes bioterapêuticos normalmente é feita por testes in vitro simulando o ambiente gastrointestinal que determina a capacidade de sobrevivência no trato digestivo. Neste trabalho, a possibilidade de extrapolação dos dados obtidos nos testes in vitro para as condições in vivo foi estudada utilizando cinco linhagens de Saccharomyces cerevisiae isoladas da floresta Atlântica brasileira. O conteúdo de trealose e a sobrevivência após a exposição a diversos estresses fisiológicos geralmente encontrados no trato gastrointestinal de humanos foram determinados para as cinco linhagens e os resultados comparados com a Saccharomyces boulardii, um probiótico conhecido. Esses resultados foram completados com a capacidade de colonização do trato gastrointestinal de camundongos gnotobióticos pelas leveduras. Pelos resultados obtidos, concluimos que os testes in vitro não são confirmados pelos ensaios in vivo, indicando que essa extrapolação não pode sempre ser feita