Jenny Vanessa Martinez Hernandez - Academia.edu (original) (raw)
Papers by Jenny Vanessa Martinez Hernandez
Lund University Faculty of Medicine Doctoral Dissertation Series, 2013
Urinary tract infections (UTIs) present an interesting and relevant model for studying microbial ... more Urinary tract infections (UTIs) present an interesting and relevant model for studying microbial adaptation. After establishing significant numbers, the bacteria either cause severe disease, or an asymptomatic carrier state resembling the normal flora at other mucosal sites. Patients with asymptomatic bacteriuria (ABU) are protected from re-infection if the strain that they carry outcompetes more pathogenic strains. Deliberate inoculation with the prototypic ABU strain Escherichia coli 83972 has therefore been developed and clinically proven to protect against recurrent UTI. To define the host influence on bacterial adaptation during long term E. coli 83972 ABU, we collected sequential isolates from patients that had been inoculated with E. coli 83972 and established stable bacteriuria. The isolates acquired several host-specific mutations, demonstrating that E. coli 83972 adapts to the individual host. Each host provided a unique niche, which was demonstrated by significant variations of mucosal host response parameters between patients. Variation in the host response to ABU has lead to uncertainty about the use of host response parameters as a basis for diagnostic and therapeutic decisions. In 23 patients, the host response to E. coli 83972 was accompanied by a low but host-specific increase in neutrophil chemotaxis but IL-6 levels did not increase. To define the effects of genetic variation on the urine proteomic host response, patients were genotyped for polymorphisms that have been linked to susceptibility to ABU and urine samples from the patients were screened for 31 immune markers. The genetic polymorphisms in the interferon regulatory factor 3 (IRF3) and Toll-like receptor 4 (TLR4) promoter had a significant impact on the magnitude of the host response during E. coli 83972 ABU, which consisted of mainly innate immune mediators. The transcriptional host response to ABU has not been examined in humans. To examine if ABU strains affect uroepithelial and leukocyte human host gene expression, we analyzed peripheral blood leukocytes from patients colonized by E. coli 83972 and uroepithelial cells stimulated with the same strain. E. coli 83972 inhibited RNA Polymerase II phosphorylation and suppressed pathogen-specific pathways in both systemic leukocytes and uroepithelial. We show that ABU is an active rather than passive process, and present a theory that basal transcriptional suppression may be a general mechanism used by commensal strains to modulate host gene expression. We further examined the kinetics of the local and systemic host response during ABU and the effect of type 1 and P fimbrial adhesion on the transcriptional signature. All patients inoculated with E. coli 83972pap activated the Interferon signaling pathway. Two patients inoculated with E. coli 83972fim downregulated Natural Killer cell signaling. Our results provide direct molecular evidence of host specific evolution of bacterial genomes as well as transcriptomic alterations in the host during ABU.
In the following work, we describe the ways of how to improve some tools taken of the Lean Manage... more In the following work, we describe the ways of how to improve some tools taken of the Lean Management philosophy in a company which sells and imports wools and fibers of them; with the purpose of quickly eradicate leftovers. Among them, the excessive number of items inventoried, increase the productivity, and improve the company as well. Lean Management is a series of techniques and improvement tools with the purpose of reduce dramatically the remnants that are the results of repetitive processes, unwarranted items inventoried, unnecessary movements or dead times, among others. With the improvement of this philosophy we can obtain a positive progress on the company and thus a better quality of service. In the present work, it is explained how these tools could help us how to eradicate scraps; or at least, decrease dramatically and how the company will improve.
Dedico este trabajo a mis padres, a mis hijas, a Gustavo por su amor y comprensión quienes supier... more Dedico este trabajo a mis padres, a mis hijas, a Gustavo por su amor y comprensión quienes supieron apoyarme para seguir adelante y culminar este reto tan importante y con éxitos mis objetivos propuestos. iii AGRADECIMIENTO Un agradecimiento muy especial al Dr. Patricio Baquero MSc. Quien en su momento determinado me instruyo para poder culminar con gran éxito este presente trabajo. Y un profundo agradecimiento nuevamente a mis padres a mi hermano y cuñada quienes a la distancia fueron la base sólida de armonía y amor, a la
La mastitis bovina es uno de los problemas de salud animal que más afecta la calidad de la leche ... more La mastitis bovina es uno de los problemas de salud animal que más afecta la calidad de la leche y la competitividad de la cadena láctea. El principal agente causal de la mastitis bovina contagiosa es Staphylococcus aureus, que generalmente produce infecciones de larga duración. La presencia del microorganismo y sus toxinas en la leche y derivados lácteos adicionalmente constituye un problema de salud pública, además de su relación con el uso incorrecto e indiscriminado de antimicrobianos que contaminan la leche e inhiben la fermentación de los cultivos bacterianos que se utilizan en la fabricación de derivados lácteos. S. aureus resistente a meticilina (MRSA), es un tipo de estafilococo resistente a todos los antimicrobianos β-lactámicos y a sus derivados. El gen responsable de dicha resistencia es el mecA, que será detectado en cepas aisladas de leche cruda provenientes de fincas de Cundinamarca, mediante la técnica de PCR.
RESUMEN La presente investigación está enfocada a evaluar la calidad higiénica y aptitud industri... more RESUMEN La presente investigación está enfocada a evaluar la calidad higiénica y aptitud industrial de la leche cruda de tanques y cantinas provenientes de 16 Fincas del Departamento de Cundinamarca, para esto se analizaron 96 muestras de leches de tanques con base en el protocolo establecido por el National Mastitis Council (NMC) de los Estados Unidos durante 6 meses. A estas muestras se les realizaron recuentos para mesófilos aerobios (SPC), Recuento de coliformes totales (CC), prueba de incubación preliminar (PIC), recuento pospasteurización en laboratorio (LPC), también se realizaron aislamientos de los microorganismos presentes en la leche proveniente de los tanques. Según los resultados, tan solo el 43,75% de las fincas entre las que lograron alcanzar estándares de calidad higiénica para recuentos de UFC/mL encontrándose por debajo del rango 175.000 – 200.000 UFC/mL. Los recuentos para evaluar microorganismos psicrotróficos y termoduricos (PIC y LPC), se encontraron altos en l...
La mastitis bovina es una enfermedad de distribución mundial, de alto impacto para la industria l... more La mastitis bovina es una enfermedad de distribución mundial, de alto impacto para la industria láctea; la identificación precisa y oportuna de géneros y especies de la familia Streptococcaceae es de gran importancia debido a que proporciona un mapa del estado sanitario del hato, según el origen de la enfermedad. El objetivo de este estudio fue realizar la identificación completa de aislamientos de la familia Streptococcaceae provenientes de muestras de leche de vacas con mastitis mediante el uso de un método convencional y uno automatizado, el Sistema Vitek ®. Se identificaron las tres especies ya reportadas por otros autores en Colombia Streptococcus agalactiae, Streptococcus uberis, Streptococcus dysgalactiae y otras ocho especies del mismo genero, antes no reportados en el país, pero si en otros países, como agentes causales de la mastitis: La mastitis bovina es una inflamación de la glándula mamaria, debida a infección o traumatismo(1). Esta enfermedad es de distribución mundia...
PLoS ONE, 2011
The severity of urinary tract infection (UTI) reflects the quality and magnitude of the host resp... more The severity of urinary tract infection (UTI) reflects the quality and magnitude of the host response. While strong local and systemic innate immune activation occurs in patients with acute pyelonephritis, the response to asymptomatic bacteriuria (ABU) is low. The immune response repertoire in ABU has not been characterized, due to the inherent problem to distinguish bacterial differences from host-determined variation. In this study, we investigated the host response to ABU and genetic variants affecting innate immune signaling and UTI susceptibility. Patients were subjected to therapeutic urinary tract inoculation with E. coli 83972 to ensure that they were exposed to the same E. coli strain. The innate immune response repertoire was characterized in urine samples, collected from each patient before and after inoculation with bacteria or PBS, if during the placebo arm of the study. Long-term E. coli 83972 ABU was established in 23 participants, who were followed for up to twelve months and the innate immune response was quantified in 233 urine samples. Neutrophil numbers increased in all but two patients and in an extended urine cytokine/chemokine analysis (31 proteins), the chemoattractants IL-8 and GRO-a, RANTES, Eotaxin-1 and MCP-1, the T cell chemoattractant and antibacterial peptide IP-10, inflammatory regulators IL-1-a and sIL-1RA and the T lymphocyte/dendritic cell product sIL-2Ra were detected and variably increased, compared to sterile samples. IL-6, which is associated with symptomatic UTI, remained low and numerous specific immune mediators were not detected. The patients were also genotyped for UTI-associated IRF3 and TLR4 promoter polymorphisms. Patients with ABU associated TLR4 polymorphisms had low neutrophil numbers, IL-6, IP-10, MCP-1 and sIL-2Ra concentrations. Patients with the ABU-associated IRF3 genotype had lower neutrophils, IL-6 and MCP-1 responses than the remaining group. The results suggest that the host-specific, low immune response to ABU mainly includes innate immune mediators and that host genetics directly influence the magnitude of this response.
Biomédica : revista del Instituto Nacional de Salud, 2003
Antigen profiles were described for Giardia duodenalis cysts and trophozoites that are recognized... more Antigen profiles were described for Giardia duodenalis cysts and trophozoites that are recognized by IgG and its anti-G. dudodenalis subclasses (IgG1, IgG2, IgG3, IgG4). Antigens were identified by Western blot from G. duodenalis cyst and trophozoite isolates. Cysts and trophozoites were each subjected to protein separation by SDS-PAGE. The proteins were then transferred to nitrocellulose membranes by electroimmunoblot, and their antigenicity was determined by exposing them to sera from patients with confirmed diagnosis of G. duodenalis infection. The antigen-antibody reaction was revealed by specific alkaline phosphatase antibody conjugates against IgG, IgG2, IgG3, IgG4: bands were visualized by addition of the substrate 5-bromo-4-chloro-3-indolyl-phosphate and the stain nitro blue tetrazolium. The bands were read and analyzed by linear regression using Quantity One software. Thirty two antigens were simultaneously recognized by total IgG anti-G. duodenalis in the cyst and trophozo...
Biomédica, 2013
Introducción. En Colombia, la práctica de lactancia exclusiva y total es inadecuada y de corta du... more Introducción. En Colombia, la práctica de lactancia exclusiva y total es inadecuada y de corta duración, especialmente en mujeres adolescentes. Teniendo en cuenta que las adolescentes constituyen un grupo social con sus propios estilos de vida, es necesario conocer cuáles son los significados que construyen alrededor de la lactancia y cuáles son las características de su experiencia de lactar para identificar aspectos que limitan o facilitan esta práctica, generando conocimientos para mejorar las estrategias de promoción de la lactancia. Objetivo. Caracterizar la experiencia de lactancia materna en adolescentes en período de lactancia, e identificar aspectos positivos, limitaciones y necesidades sentidas desde su propia perspectiva. Materiales y métodos. Se trata de un estudio cualitativo con enfoque fenomenológico. Se hicieron 24 entrevistas y se conformaron tres grupos de mujeres adolescentes en diferentes períodos posparto. La información se recolectó en Bogotá con mujeres que participaban en un programa de la Secretaría Distrital de Integración Social. El proceso de sistematización se desarrolló paralelamente con el proceso de análisis. Este proceso comprendió las relaciones entre categorías y las redes que se forman entre ellas. Resultados. Las adolescentes no alimentan sus hijos con lactancia materna exclusivamente y refieren varias dificultades en el acto de lactar. La alimentación complementaria incluye alimentos no naturales. La maternidad y la lactancia no son acordes con la percepción de ser adolescente. Conclusiones. Las adolescentes reconocen los beneficios que ofrece la lactancia materna para sus hijos y para ellas; sin embargo, su experiencia de lactancia materna difiere de las recomendaciones para lograr una lactancia materna exclusiva y una alimentación complementaria saludable. Entre las causas identificadas, sobresale la ausencia de acompañamiento acertado y oportuno.
Proceedings of the National Academy of Sciences, 2005
Despite the increasing availability of genome sequences from many human pathogens, the production... more Despite the increasing availability of genome sequences from many human pathogens, the production of complete proteomes remains at a bottleneck. To address this need, a high-throughput PCR recombination cloning and expression platform has been developed that allows hundreds of genes to be batch-processed by using ordinary laboratory procedures without robotics. The method relies on high-throughput amplification of each predicted ORF by using gene specific primers, followed by in vivo homologous recombination into a T7 expression vector. The proteins are expressed in an Escherichia coli -based cell-free in vitro transcription/translation system, and the crude reactions containing expressed proteins are printed directly onto nitrocellulose microarrays without purification. The protein microarrays are useful for determining the complete antigen-specific humoral immune-response profile from vaccinated or infected humans and animals. The system was verified by cloning, expressing, and pr...
Journal of Virology, 2007
Modified vaccinia virus Ankara (MVA) is a highly attenuated vaccinia virus that is under consider... more Modified vaccinia virus Ankara (MVA) is a highly attenuated vaccinia virus that is under consideration as an alternative to the conventional smallpox vaccine Dryvax. MVA was attenuated by extensive passage of vaccinia virus Ankara in chicken embryo fibroblasts. Several immunomodulatory genes and genes that influence host range are deleted or mutated, and replication is aborted in the late stage of infection in most nonavian cells. The effect of these mutations on immunogenicity is not well understood. Since the structural genes appear to be intact in MVA, it is hypothesized that critical targets for antibody neutralization have been retained. To test this, we probed microarrays of the Western Reserve (WR) proteome with sera from humans and macaques after MVA and Dryvax vaccination. As most protein sequences of MVA are 97 to 99% identical to those of other vaccinia virus strains, extensive binding cross-reactivity is expected, except for those deleted or truncated. Despite different ...
Journal of Virology, 2005
The smallpox vaccine is the prototypic vaccine, yet the viral targets critical for vaccine-mediat... more The smallpox vaccine is the prototypic vaccine, yet the viral targets critical for vaccine-mediated protection remain unclear in humans. We have produced protein microarrays of a near-complete vaccinia proteome and used them to determine the major antigen specificities of the human humoral immune response to the smallpox vaccine (Dryvax). H3L, an intracellular mature virion envelope protein, was consistently recognized by high-titer antibodies in the majority of human donors, particularly after secondary immunization. We then focused on examining H3L as a valuable human antibody target. Purified human anti-H3L antibodies exhibited substantial vaccinia virus-neutralizing activity in vitro (50% plaque reduction neutralization test [PRNT 50 ] = 44 μg/ml). Mice also make an immunodominant antibody response to H3L after vaccination with vaccinia virus, as determined by vaccinia virus protein microarray. Mice were immunized with recombinant H3L protein to examine H3L-specific antibody res...
The Journal of Urology, 2014
Purpose: Asymptomatic bacteriuria established by intravesical inoculation of Escherichia coli 839... more Purpose: Asymptomatic bacteriuria established by intravesical inoculation of Escherichia coli 83972 is protective in patients with recurrent urinary tract infections. In this randomized, controlled crossover study a total of 3 symptomatic urinary tract infection episodes developed in 2 patients while they carried E. coli 83972. We examined whether virulence reacquisition by symptom isolates may account for the switch from asymptomatic bacteriuria to symptomatic urinary tract infection. Materials and Methods: We used E. coli 83972 re-isolates from 2 patients in a prospective study and from another 2 in whom symptoms developed after study completion. We phylogenetically classified the re-isolates, and identified the genomic restriction patterns and gene expression profiles as well as virulence gene structure and phenotypes. In vivo virulence was examined in the murine urinary tract infection model. Results: The fim, pap, foc, hlyA, fyuA, iuc, iroN, kpsMT K5 and malX genotypes of the symptomatic re-isolates remained unchanged. Bacterial gene expression profiles of flagellated symptomatic re-isolates were unique to each host, providing no evidence of common deregulation. Symptomatic isolates did not differ in virulence from the wild-type strain, as defined in the murine urinary tract infection model by persistence, symptoms or innate immune activation. Conclusions: The switch from asymptomatic E. coli 83972 carriage to symptomatic urinary tract infection was not explained by reversion to a functional virulence gene repertoire.
Journal of Clinical Investigation, 2013
The normal flora furnishes the host with ecological barriers that prevent pathogen attack while m... more The normal flora furnishes the host with ecological barriers that prevent pathogen attack while maintaining tissue homeostasis. Urinary tract infections (UTIs) constitute a highly relevant model of microbial adaptation in which some patients infected with Escherichia coli develop acute pyelonephritis, while other patients with bacteriuria exhibit an asymptomatic carrier state similar to bacterial commensalism. It remains unclear if the lack of destructive inflammation merely reflects low virulence or if carrier strains actively inhibit disease-associated responses in the host. Here, we identify a new mechanism of bacterial adaptation through broad suppression of RNA polymerase II-dependent (Pol II-dependent) host gene expression. Over 60% of all genes were suppressed 24 hours after human inoculation with the prototype asymptomatic bacteriuria (ABU) strain E. coli 83972, and inhibition was verified by infection of human cells. Specific repressors and activators of Pol II-dependent transcription were modified, Pol II phosphorylation was inhibited, and pathogen-specific signaling was suppressed in cell lines and inoculated patients. An increased frequency of strains inhibiting Pol II was epidemiologically verified in ABU and fecal strains compared with acute pyelonephritis, and a Pol II antagonist suppressed the disease-associated host response. These results suggest that by manipulating host gene expression, ABU strains promote tissue integrity while inhibiting pathology. Such bacterial modulation of host gene expression may be essential to sustain asymptomatic bacterial carriage by ensuring that potentially destructive immune activation will not occur. Conflict of interest: The authors have declared that no conflict of interest exists.
Lund University Faculty of Medicine Doctoral Dissertation Series, 2013
Urinary tract infections (UTIs) present an interesting and relevant model for studying microbial ... more Urinary tract infections (UTIs) present an interesting and relevant model for studying microbial adaptation. After establishing significant numbers, the bacteria either cause severe disease, or an asymptomatic carrier state resembling the normal flora at other mucosal sites. Patients with asymptomatic bacteriuria (ABU) are protected from re-infection if the strain that they carry outcompetes more pathogenic strains. Deliberate inoculation with the prototypic ABU strain Escherichia coli 83972 has therefore been developed and clinically proven to protect against recurrent UTI. To define the host influence on bacterial adaptation during long term E. coli 83972 ABU, we collected sequential isolates from patients that had been inoculated with E. coli 83972 and established stable bacteriuria. The isolates acquired several host-specific mutations, demonstrating that E. coli 83972 adapts to the individual host. Each host provided a unique niche, which was demonstrated by significant variations of mucosal host response parameters between patients. Variation in the host response to ABU has lead to uncertainty about the use of host response parameters as a basis for diagnostic and therapeutic decisions. In 23 patients, the host response to E. coli 83972 was accompanied by a low but host-specific increase in neutrophil chemotaxis but IL-6 levels did not increase. To define the effects of genetic variation on the urine proteomic host response, patients were genotyped for polymorphisms that have been linked to susceptibility to ABU and urine samples from the patients were screened for 31 immune markers. The genetic polymorphisms in the interferon regulatory factor 3 (IRF3) and Toll-like receptor 4 (TLR4) promoter had a significant impact on the magnitude of the host response during E. coli 83972 ABU, which consisted of mainly innate immune mediators. The transcriptional host response to ABU has not been examined in humans. To examine if ABU strains affect uroepithelial and leukocyte human host gene expression, we analyzed peripheral blood leukocytes from patients colonized by E. coli 83972 and uroepithelial cells stimulated with the same strain. E. coli 83972 inhibited RNA Polymerase II phosphorylation and suppressed pathogen-specific pathways in both systemic leukocytes and uroepithelial. We show that ABU is an active rather than passive process, and present a theory that basal transcriptional suppression may be a general mechanism used by commensal strains to modulate host gene expression. We further examined the kinetics of the local and systemic host response during ABU and the effect of type 1 and P fimbrial adhesion on the transcriptional signature. All patients inoculated with E. coli 83972pap activated the Interferon signaling pathway. Two patients inoculated with E. coli 83972fim downregulated Natural Killer cell signaling. Our results provide direct molecular evidence of host specific evolution of bacterial genomes as well as transcriptomic alterations in the host during ABU.
In the following work, we describe the ways of how to improve some tools taken of the Lean Manage... more In the following work, we describe the ways of how to improve some tools taken of the Lean Management philosophy in a company which sells and imports wools and fibers of them; with the purpose of quickly eradicate leftovers. Among them, the excessive number of items inventoried, increase the productivity, and improve the company as well. Lean Management is a series of techniques and improvement tools with the purpose of reduce dramatically the remnants that are the results of repetitive processes, unwarranted items inventoried, unnecessary movements or dead times, among others. With the improvement of this philosophy we can obtain a positive progress on the company and thus a better quality of service. In the present work, it is explained how these tools could help us how to eradicate scraps; or at least, decrease dramatically and how the company will improve.
Dedico este trabajo a mis padres, a mis hijas, a Gustavo por su amor y comprensión quienes supier... more Dedico este trabajo a mis padres, a mis hijas, a Gustavo por su amor y comprensión quienes supieron apoyarme para seguir adelante y culminar este reto tan importante y con éxitos mis objetivos propuestos. iii AGRADECIMIENTO Un agradecimiento muy especial al Dr. Patricio Baquero MSc. Quien en su momento determinado me instruyo para poder culminar con gran éxito este presente trabajo. Y un profundo agradecimiento nuevamente a mis padres a mi hermano y cuñada quienes a la distancia fueron la base sólida de armonía y amor, a la
La mastitis bovina es uno de los problemas de salud animal que más afecta la calidad de la leche ... more La mastitis bovina es uno de los problemas de salud animal que más afecta la calidad de la leche y la competitividad de la cadena láctea. El principal agente causal de la mastitis bovina contagiosa es Staphylococcus aureus, que generalmente produce infecciones de larga duración. La presencia del microorganismo y sus toxinas en la leche y derivados lácteos adicionalmente constituye un problema de salud pública, además de su relación con el uso incorrecto e indiscriminado de antimicrobianos que contaminan la leche e inhiben la fermentación de los cultivos bacterianos que se utilizan en la fabricación de derivados lácteos. S. aureus resistente a meticilina (MRSA), es un tipo de estafilococo resistente a todos los antimicrobianos β-lactámicos y a sus derivados. El gen responsable de dicha resistencia es el mecA, que será detectado en cepas aisladas de leche cruda provenientes de fincas de Cundinamarca, mediante la técnica de PCR.
RESUMEN La presente investigación está enfocada a evaluar la calidad higiénica y aptitud industri... more RESUMEN La presente investigación está enfocada a evaluar la calidad higiénica y aptitud industrial de la leche cruda de tanques y cantinas provenientes de 16 Fincas del Departamento de Cundinamarca, para esto se analizaron 96 muestras de leches de tanques con base en el protocolo establecido por el National Mastitis Council (NMC) de los Estados Unidos durante 6 meses. A estas muestras se les realizaron recuentos para mesófilos aerobios (SPC), Recuento de coliformes totales (CC), prueba de incubación preliminar (PIC), recuento pospasteurización en laboratorio (LPC), también se realizaron aislamientos de los microorganismos presentes en la leche proveniente de los tanques. Según los resultados, tan solo el 43,75% de las fincas entre las que lograron alcanzar estándares de calidad higiénica para recuentos de UFC/mL encontrándose por debajo del rango 175.000 – 200.000 UFC/mL. Los recuentos para evaluar microorganismos psicrotróficos y termoduricos (PIC y LPC), se encontraron altos en l...
La mastitis bovina es una enfermedad de distribución mundial, de alto impacto para la industria l... more La mastitis bovina es una enfermedad de distribución mundial, de alto impacto para la industria láctea; la identificación precisa y oportuna de géneros y especies de la familia Streptococcaceae es de gran importancia debido a que proporciona un mapa del estado sanitario del hato, según el origen de la enfermedad. El objetivo de este estudio fue realizar la identificación completa de aislamientos de la familia Streptococcaceae provenientes de muestras de leche de vacas con mastitis mediante el uso de un método convencional y uno automatizado, el Sistema Vitek ®. Se identificaron las tres especies ya reportadas por otros autores en Colombia Streptococcus agalactiae, Streptococcus uberis, Streptococcus dysgalactiae y otras ocho especies del mismo genero, antes no reportados en el país, pero si en otros países, como agentes causales de la mastitis: La mastitis bovina es una inflamación de la glándula mamaria, debida a infección o traumatismo(1). Esta enfermedad es de distribución mundia...
PLoS ONE, 2011
The severity of urinary tract infection (UTI) reflects the quality and magnitude of the host resp... more The severity of urinary tract infection (UTI) reflects the quality and magnitude of the host response. While strong local and systemic innate immune activation occurs in patients with acute pyelonephritis, the response to asymptomatic bacteriuria (ABU) is low. The immune response repertoire in ABU has not been characterized, due to the inherent problem to distinguish bacterial differences from host-determined variation. In this study, we investigated the host response to ABU and genetic variants affecting innate immune signaling and UTI susceptibility. Patients were subjected to therapeutic urinary tract inoculation with E. coli 83972 to ensure that they were exposed to the same E. coli strain. The innate immune response repertoire was characterized in urine samples, collected from each patient before and after inoculation with bacteria or PBS, if during the placebo arm of the study. Long-term E. coli 83972 ABU was established in 23 participants, who were followed for up to twelve months and the innate immune response was quantified in 233 urine samples. Neutrophil numbers increased in all but two patients and in an extended urine cytokine/chemokine analysis (31 proteins), the chemoattractants IL-8 and GRO-a, RANTES, Eotaxin-1 and MCP-1, the T cell chemoattractant and antibacterial peptide IP-10, inflammatory regulators IL-1-a and sIL-1RA and the T lymphocyte/dendritic cell product sIL-2Ra were detected and variably increased, compared to sterile samples. IL-6, which is associated with symptomatic UTI, remained low and numerous specific immune mediators were not detected. The patients were also genotyped for UTI-associated IRF3 and TLR4 promoter polymorphisms. Patients with ABU associated TLR4 polymorphisms had low neutrophil numbers, IL-6, IP-10, MCP-1 and sIL-2Ra concentrations. Patients with the ABU-associated IRF3 genotype had lower neutrophils, IL-6 and MCP-1 responses than the remaining group. The results suggest that the host-specific, low immune response to ABU mainly includes innate immune mediators and that host genetics directly influence the magnitude of this response.
Biomédica : revista del Instituto Nacional de Salud, 2003
Antigen profiles were described for Giardia duodenalis cysts and trophozoites that are recognized... more Antigen profiles were described for Giardia duodenalis cysts and trophozoites that are recognized by IgG and its anti-G. dudodenalis subclasses (IgG1, IgG2, IgG3, IgG4). Antigens were identified by Western blot from G. duodenalis cyst and trophozoite isolates. Cysts and trophozoites were each subjected to protein separation by SDS-PAGE. The proteins were then transferred to nitrocellulose membranes by electroimmunoblot, and their antigenicity was determined by exposing them to sera from patients with confirmed diagnosis of G. duodenalis infection. The antigen-antibody reaction was revealed by specific alkaline phosphatase antibody conjugates against IgG, IgG2, IgG3, IgG4: bands were visualized by addition of the substrate 5-bromo-4-chloro-3-indolyl-phosphate and the stain nitro blue tetrazolium. The bands were read and analyzed by linear regression using Quantity One software. Thirty two antigens were simultaneously recognized by total IgG anti-G. duodenalis in the cyst and trophozo...
Biomédica, 2013
Introducción. En Colombia, la práctica de lactancia exclusiva y total es inadecuada y de corta du... more Introducción. En Colombia, la práctica de lactancia exclusiva y total es inadecuada y de corta duración, especialmente en mujeres adolescentes. Teniendo en cuenta que las adolescentes constituyen un grupo social con sus propios estilos de vida, es necesario conocer cuáles son los significados que construyen alrededor de la lactancia y cuáles son las características de su experiencia de lactar para identificar aspectos que limitan o facilitan esta práctica, generando conocimientos para mejorar las estrategias de promoción de la lactancia. Objetivo. Caracterizar la experiencia de lactancia materna en adolescentes en período de lactancia, e identificar aspectos positivos, limitaciones y necesidades sentidas desde su propia perspectiva. Materiales y métodos. Se trata de un estudio cualitativo con enfoque fenomenológico. Se hicieron 24 entrevistas y se conformaron tres grupos de mujeres adolescentes en diferentes períodos posparto. La información se recolectó en Bogotá con mujeres que participaban en un programa de la Secretaría Distrital de Integración Social. El proceso de sistematización se desarrolló paralelamente con el proceso de análisis. Este proceso comprendió las relaciones entre categorías y las redes que se forman entre ellas. Resultados. Las adolescentes no alimentan sus hijos con lactancia materna exclusivamente y refieren varias dificultades en el acto de lactar. La alimentación complementaria incluye alimentos no naturales. La maternidad y la lactancia no son acordes con la percepción de ser adolescente. Conclusiones. Las adolescentes reconocen los beneficios que ofrece la lactancia materna para sus hijos y para ellas; sin embargo, su experiencia de lactancia materna difiere de las recomendaciones para lograr una lactancia materna exclusiva y una alimentación complementaria saludable. Entre las causas identificadas, sobresale la ausencia de acompañamiento acertado y oportuno.
Proceedings of the National Academy of Sciences, 2005
Despite the increasing availability of genome sequences from many human pathogens, the production... more Despite the increasing availability of genome sequences from many human pathogens, the production of complete proteomes remains at a bottleneck. To address this need, a high-throughput PCR recombination cloning and expression platform has been developed that allows hundreds of genes to be batch-processed by using ordinary laboratory procedures without robotics. The method relies on high-throughput amplification of each predicted ORF by using gene specific primers, followed by in vivo homologous recombination into a T7 expression vector. The proteins are expressed in an Escherichia coli -based cell-free in vitro transcription/translation system, and the crude reactions containing expressed proteins are printed directly onto nitrocellulose microarrays without purification. The protein microarrays are useful for determining the complete antigen-specific humoral immune-response profile from vaccinated or infected humans and animals. The system was verified by cloning, expressing, and pr...
Journal of Virology, 2007
Modified vaccinia virus Ankara (MVA) is a highly attenuated vaccinia virus that is under consider... more Modified vaccinia virus Ankara (MVA) is a highly attenuated vaccinia virus that is under consideration as an alternative to the conventional smallpox vaccine Dryvax. MVA was attenuated by extensive passage of vaccinia virus Ankara in chicken embryo fibroblasts. Several immunomodulatory genes and genes that influence host range are deleted or mutated, and replication is aborted in the late stage of infection in most nonavian cells. The effect of these mutations on immunogenicity is not well understood. Since the structural genes appear to be intact in MVA, it is hypothesized that critical targets for antibody neutralization have been retained. To test this, we probed microarrays of the Western Reserve (WR) proteome with sera from humans and macaques after MVA and Dryvax vaccination. As most protein sequences of MVA are 97 to 99% identical to those of other vaccinia virus strains, extensive binding cross-reactivity is expected, except for those deleted or truncated. Despite different ...
Journal of Virology, 2005
The smallpox vaccine is the prototypic vaccine, yet the viral targets critical for vaccine-mediat... more The smallpox vaccine is the prototypic vaccine, yet the viral targets critical for vaccine-mediated protection remain unclear in humans. We have produced protein microarrays of a near-complete vaccinia proteome and used them to determine the major antigen specificities of the human humoral immune response to the smallpox vaccine (Dryvax). H3L, an intracellular mature virion envelope protein, was consistently recognized by high-titer antibodies in the majority of human donors, particularly after secondary immunization. We then focused on examining H3L as a valuable human antibody target. Purified human anti-H3L antibodies exhibited substantial vaccinia virus-neutralizing activity in vitro (50% plaque reduction neutralization test [PRNT 50 ] = 44 μg/ml). Mice also make an immunodominant antibody response to H3L after vaccination with vaccinia virus, as determined by vaccinia virus protein microarray. Mice were immunized with recombinant H3L protein to examine H3L-specific antibody res...
The Journal of Urology, 2014
Purpose: Asymptomatic bacteriuria established by intravesical inoculation of Escherichia coli 839... more Purpose: Asymptomatic bacteriuria established by intravesical inoculation of Escherichia coli 83972 is protective in patients with recurrent urinary tract infections. In this randomized, controlled crossover study a total of 3 symptomatic urinary tract infection episodes developed in 2 patients while they carried E. coli 83972. We examined whether virulence reacquisition by symptom isolates may account for the switch from asymptomatic bacteriuria to symptomatic urinary tract infection. Materials and Methods: We used E. coli 83972 re-isolates from 2 patients in a prospective study and from another 2 in whom symptoms developed after study completion. We phylogenetically classified the re-isolates, and identified the genomic restriction patterns and gene expression profiles as well as virulence gene structure and phenotypes. In vivo virulence was examined in the murine urinary tract infection model. Results: The fim, pap, foc, hlyA, fyuA, iuc, iroN, kpsMT K5 and malX genotypes of the symptomatic re-isolates remained unchanged. Bacterial gene expression profiles of flagellated symptomatic re-isolates were unique to each host, providing no evidence of common deregulation. Symptomatic isolates did not differ in virulence from the wild-type strain, as defined in the murine urinary tract infection model by persistence, symptoms or innate immune activation. Conclusions: The switch from asymptomatic E. coli 83972 carriage to symptomatic urinary tract infection was not explained by reversion to a functional virulence gene repertoire.
Journal of Clinical Investigation, 2013
The normal flora furnishes the host with ecological barriers that prevent pathogen attack while m... more The normal flora furnishes the host with ecological barriers that prevent pathogen attack while maintaining tissue homeostasis. Urinary tract infections (UTIs) constitute a highly relevant model of microbial adaptation in which some patients infected with Escherichia coli develop acute pyelonephritis, while other patients with bacteriuria exhibit an asymptomatic carrier state similar to bacterial commensalism. It remains unclear if the lack of destructive inflammation merely reflects low virulence or if carrier strains actively inhibit disease-associated responses in the host. Here, we identify a new mechanism of bacterial adaptation through broad suppression of RNA polymerase II-dependent (Pol II-dependent) host gene expression. Over 60% of all genes were suppressed 24 hours after human inoculation with the prototype asymptomatic bacteriuria (ABU) strain E. coli 83972, and inhibition was verified by infection of human cells. Specific repressors and activators of Pol II-dependent transcription were modified, Pol II phosphorylation was inhibited, and pathogen-specific signaling was suppressed in cell lines and inoculated patients. An increased frequency of strains inhibiting Pol II was epidemiologically verified in ABU and fecal strains compared with acute pyelonephritis, and a Pol II antagonist suppressed the disease-associated host response. These results suggest that by manipulating host gene expression, ABU strains promote tissue integrity while inhibiting pathology. Such bacterial modulation of host gene expression may be essential to sustain asymptomatic bacterial carriage by ensuring that potentially destructive immune activation will not occur. Conflict of interest: The authors have declared that no conflict of interest exists.