RESUMEN
The complex microbial population residing in the human gastrointestinal tract consists of commensal, potential pathogenic and beneficial species, which are probably perceived differently by the host and consequently could be expected to trigger specific transcriptional responses. Here, we provide a comparative analysis of the global in vitro transcriptional response of human intestinal epithelial cells to Lactobacillus acidophilus NCFM™, Lactobacillus salivarius Ls-33, Bifidobacterium animalis subsp. lactis 420, and enterohaemorrhagic Escherichia coli O157:H7 (EHEC). Interestingly, L. salivarius Ls-33 DCE-induced changes were overall more similar to those of B. lactis 420 than to L. acidophilus NCFM™, which is consistent with previously observed in vivo immunomodulation properties. In the gene ontology and pathway analyses both specific and unspecific changes were observed. Common to all was the regulation of apoptosis and adipogenesis, and lipid-metabolism related regulation by the probiotics. Specific changes such as regulation of cell-cell adhesion by B. lactis 420, superoxide metabolism by L. salivarius Ls-33, and regulation of MAPK pathway by L. acidophilus NCFM™ were noted. Furthermore, fundamental differences were observed between the pathogenic and probiotic treatments in the Toll-like receptor pathway, especially for adapter molecules with a lowered level of transcriptional activation of MyD88, TRIF, IRAK1 and TRAF6 by probiotics compared to EHEC. The results in this study provide insights into the relationship between probiotics and human intestinal epithelial cells, notably with regard to strain-specific responses, and highlight the differences between transcriptional responses to pathogenic and probiotic bacteria.
Asunto(s)
Bifidobacterium/fisiología , Células Epiteliales/metabolismo , Escherichia coli/fisiología , Regulación de la Expresión Génica , Intestinos/citología , Lactobacillus/fisiología , Línea Celular , Células Epiteliales/efectos de los fármacos , Células Epiteliales/microbiología , Perfilación de la Expresión Génica , Regulación de la Expresión Génica/efectos de los fármacos , Humanos , Mucosa Intestinal/metabolismo , Intestinos/efectos de los fármacos , Intestinos/microbiología , Modelos Biológicos , Probióticos/farmacología , Transducción de Señal , Activación Transcripcional/efectos de los fármacosRESUMEN
To evaluate the potential of indigestible oligosaccharides (OS) to serve as "dietary fiber-like" ingredients, it is necessary to determine their extent of indigestibility. In vitro fermentation characteristics of two novel OS, alpha-glucooligosaccharides (GOS) and a maltodextrin-like OS (MD), were compared to those of fructooligosaccharides (FOS), gum arabic (GA), guar gum (GG) and guar hydrolysate (GH). Total short-chain fatty acid (SCFA) production (micromol/g dry matter) as a result of MD fermentation was higher initially compared with GA (P<0.01), but GA was more extensively fermented at 24 h (P<0.01). Total SCFA production for GOS was similar to that for FOS, GG, GH and GA. In the second experiment, GOS and MD were added at 6% to an enteral formula control diet (Control) and fed to ileal-cannulated dogs in a 3x3 replicated Latin-square design. Ileal digestibility of glucose was lower (P<0.05) and carbohydrate (CHO) numerically lower (P = 0.08) for both GOS and MD compared with the Control. Total tract digestibility of CHO and glucose was lower only for MD (P<0.01) compared with the Control. Total fecal weights were higher (P<0.01) for both GOS and MD treatments. Fecal concentration of bifidobacteria was numerically increased by GOS and MD supplementation (P = 0.13 and 0.23, respectively). Thus, GOS and MD are indigestible yet fermentable OS, and may act as "dietary fiber-like" ingredients.
Asunto(s)
Bacteroides/efectos de los fármacos , Dieta , Digestión , Ácidos Grasos Volátiles/biosíntesis , Heces/microbiología , Intestinos/microbiología , Oligosacáridos/farmacología , Animales , Cromatografía Líquida de Alta Presión , Digestión/efectos de los fármacos , Perros , Nutrición Enteral , Femenino , Fermentación , Glucosa/metabolismo , Concentración de Iones de Hidrógeno , Mucosa Intestinal/metabolismo , Oligosacáridos/administración & dosificación , Especificidad por SustratoRESUMEN
Acid adaptation of Salmonella typhimurium at a pH of 5.0 to 5.8 for one to two cell doublings resulted in marked sensitization of the pathogen to halogen-based sanitizers including chlorine (hypochlorous acid) and iodine. Acid-adapted S. typhimurium was more resistant to an anionic acid sanitizer than was its nonadapted counterpart. A nonselective plating medium of tryptose phosphate agar plus 1% pyruvate was used throughout the study to help recover chemically stressed cells. Mechanisms of HOCl-mediated inactivation of acid-adapted and nonadapted salmonellae were investigated. Hypochlorous acid oxidized a higher percentage of cell surface sulfhydryl groups in acid-adapted cells than in nonadapted cells, and sulfhydryl oxidation was correlated with cell inactivation. HOCl caused severe metabolic disruptions in acid-adapted and nonadapted S. typhimurium, such as respiratory loss and inability to restore the adenylate energy charge from a nutrient-starved state. Sensitization of S. typhimurium to hypochlorous acid by acid adaptation also involved increased permeability of the cell surface because nonadapted cells treated with EDTA became sensitized. The results of this study establish that acid-adapted S. typhimurium cells are highly sensitized to HOCl oxidation and that inactivation by HOCl involves changes in membrane permeability, inability to maintain or restore energy charge, and probably oxidation of essential cellular components. This study provides a basis for improved practical technologies to inactivate Salmonella and implies that acid pretreatment of food plant environments may increase the efficacy of halogen sanitizers.
Asunto(s)
Ácidos/farmacología , Adaptación Biológica , Desinfectantes/farmacología , Ácido Hipocloroso/farmacología , Salmonella typhimurium/fisiología , Permeabilidad de la Membrana Celular/efectos de los fármacos , Relación Dosis-Respuesta a Droga , Interacciones Farmacológicas , Metabolismo Energético/efectos de los fármacos , Inhibidores Enzimáticos/farmacología , Concentración de Iones de Hidrógeno , Yodo/farmacología , Pruebas de Sensibilidad Microbiana , Oxidorreductasas/efectos de los fármacos , Salmonella typhimurium/efectos de los fármacos , Compuestos de SulfhidriloRESUMEN
Escherichia coli O157:H7 was adapted to acid by culturing for one to two doublings at pH 5.0. Acid-adapted cells had an increased resistance to lactic acid, survived better than nonadapted cells during a sausage fermentation, and showed enhanced survival in shredded dry salami (pH 5.0) and apple cider (pH 3.4). Acid adaptation is important for the survival of E. coli O157:H7 in acidic foods and should be considered a prerequisite for inocula used in food challenge studies.
Asunto(s)
Escherichia coli/fisiología , Microbiología de Alimentos , Ácidos , Adaptación Fisiológica , Animales , Bebidas/microbiología , Diarrea/etiología , Escherichia coli/efectos de los fármacos , Escherichia coli/patogenicidad , Infecciones por Escherichia coli/etiología , Fermentación , Enfermedades Transmitidas por los Alimentos/etiología , Frutas , Humanos , Concentración de Iones de Hidrógeno , Lactatos/farmacología , Ácido Láctico , Productos de la Carne/microbiologíaRESUMEN
Listeria monocytogenes was highly resistant to hen egg white lysozyme in whole milk but was sensitive in media and in phosphate buffer. Methods to sensitize the pathogen to lysozyme in milk were investigated. Treatment of whole milk by cation exchange to remove minerals, particularly Ca2+ and Mg2+, slightly promoted inactivation of L. monocytogenes by lysozyme at 4 degrees C over a period of 6 days. Heat treatment (62.5 degrees C for 15 s) strongly sensitized L. monocytogenes to lysozyme in demineralized milk and in MES [2-(N-morpholino)ethanesulfonic acid] buffer. Addition of Ca2+ or Mg2+ to the demineralized milk restored resistance to lysozyme. Cells were more rapidly heat inactivated at 55 degrees C in demineralized milk containing lysozyme, and addition of Ca2+ to the demineralized milk restored the resistance to heat. The results indicate that minerals or mineral-associated components protect L. monocytogenes from inactivation by lysozyme and heat in milk, probably by increasing cell surface stability. The heat treatment of foods containing added lysozyme can probably play a significant role in producing microbiologically safe foods.
Asunto(s)
Listeria monocytogenes/efectos de los fármacos , Leche/enzimología , Leche/microbiología , Muramidasa/farmacología , Animales , Tampones (Química) , Pollos , Farmacorresistencia Microbiana , Ácido Edético/farmacología , Calor , Listeria monocytogenes/ultraestructura , Metales/farmacología , Microscopía Electrónica de RastreoRESUMEN
The relationship of acid adaptation to tolerance of other environmental stresses was examined in Salmonella typhimurium. S. typhimurium was adapted to acid by exposing the cells to mildly acidic conditions (pH 5.8) for one to two cell doublings. Acid-adapted cells were found to have increased tolerance towards various stresses including heat, salt, an activated lactoperoxidase system, and the surface-active agents crystal violet and polymyxin B. Acid adaptation increased cell surface hydrophobicity. Specific outer membrane proteins were induced by acid adaptation, but the lipopolysaccharide component appeared to be unaltered. These results show that acid adaptation alters cellular resistance to a variety of environmental stresses. The mechanism of acid-induced cross-protection involved changes in cell surface properties in addition to the known enhancement of intracellular pH homeostasis.
Asunto(s)
Adaptación Fisiológica , Proteínas de la Membrana Bacteriana Externa/biosíntesis , Calor , Concentración de Iones de Hidrógeno , Lactoperoxidasa/farmacología , Salmonella typhimurium/fisiología , Cloruro de Sodio/farmacología , Violeta de Genciana/farmacología , Polimixina B/farmacología , Salmonella typhimurium/efectos de los fármacos , Salmonella typhimurium/crecimiento & desarrolloRESUMEN
Salmonella typhimurium was adapted to acid by exposure to hydrochloric acid at pH 5.8 for one to two doublings. Acid-adapted cells had increased resistance to inactivation by organic acids commonly present in cheese, including lactic, propionic, and acetic acids. Recovery of cells during the treatment with organic acids was increased 1,000-fold by inclusion of 0.1% sodium pyruvate in the recovery medium. Acid-adapted S. typhimurium cells survived better than nonadapted cells during a milk fermentation by a lactic acid culture. Acid-adapted cells also showed enhanced survival over a period of two months in cheddar, Swiss, and mozzarella cheeses kept at 5 degrees C. Acid adaptation was found in Salmonella spp., including Salmonella enteritidis, Salmonella choleraesuis subsp. choleraesuis serotype heidelberg, and Salmonella choleraesuis subsp. choleraesuis serotype javiana, associated with food poisoning. These observations support the theory that acid adaptation is an important survival mechanism enabling Salmonella spp. to persist in fermented dairy products and possibly other acidic food products.
Asunto(s)
Microbiología de Alimentos , Salmonella/aislamiento & purificación , Adaptación Fisiológica , Animales , Queso/microbiología , Fermentación , Enfermedades Transmitidas por los Alimentos/microbiología , Humanos , Concentración de Iones de Hidrógeno , Leche , Salmonella/crecimiento & desarrollo , Salmonella/fisiologíaRESUMEN
Of the seven amino acids required by Clostridium botulinum type E, tryptophan is the most essential and may provide the cell with nitrogen. The addition of excess tryptophan (10-20 mM) or other nitrogenous nutrients to minimal growth medium markedly decreased toxin formation but did not affect growth in C. botulinum type E. On the other hand, the addition of an enzymatic digest of casein (NZ Case) stimulated toxin formation and overcame repression by tryptophan. Immunoblots of proteins in culture fluids using antibodies to type E toxin indicated that tryptophan-repressed cultures produced less neurotoxin protein. Inhibitors of neurotoxin did not accumulate in cultures grown in minimal medium supplemented with high tryptophan. The results suggest that tryptophan availability in foods or in the intestine may be important for toxin formation by C. botulinum type E.