ASTMH 64th Annual Meeting. October 25-29, 2015 Philadelphia Marriott Downtown Philadelphia, Pennsylvania USA / Bartonella bacilliformis is the aetiological agent of Carrion's disease, an overlooked illness with a lethal febrile stage and a benign warty phase. Its endemic in Andean areas, mainly affecting Peru, but also reported in Ecuador, Colombia, Bolivia and Chile.

ASTMH 64th Annual Meeting. October 25-29, 2015 Philadelphia Marriott Downtown Philadelphia, Pennsylvania USA / The bacteria Bartonella bacilliformis is the etiological agent of Carrion’s disease, which is a neglected poverty-related disease, affecting Mountain Andean valleys of Peru, Colombia and Ecuador. This disease, in absence of treatment presents a high mortality during the acute phase, called Oroya’s Fever. The second phase is characterized by the development of dermal eruptions, known as “Peruvian wart”. This bacterium is a fastidious slow growing microorganism, being difficult and cumbersome to culture and isolate from clinical sources. Then, the available data about phylogenetic relationship in clinical samples are really scarce, but suggesting high variability. The aim of the study was to perform direct blood analysis of B. bacilliformis Multi Locus Sequence Typing (MLST), a genotyping tool, in patients with Oroya fever during an outbreak. The present study demonstrate that the direct blood MLST PCR is a technique useful in the phylogenic characterization of this fastidious microorganism endemic from Andean regions. In this study, we demonstrate that the outbreak of Oroya’s fever was caused by closely related Sequence Typing (ST) microorganisms and, additionally, new STs have been described.

El estudio fue apoyado por el Programa Nacional de Innovación para la Competitividad y Productividad (Innovar Perú). / FONDO: Bartonella bacilliformis es el agente causante de la enfermedad de Carrion, una enfermedad descuidado con tasas de mortalidad de 40 a 85% en ausencia de tratamiento. La falta de una técnica de diagnóstico para superar un mal diagnóstico y el tratamiento de portadores asintomáticos es de nota. Este estudio tuvo como objetivo identificar nuevos candidatos B. bacilliformis antigénicos que podrían dar lugar a una nueva herramienta de diagnóstico capaz de ser implementado en las zonas rurales endémicas.

In blood banks the sought for a series of relevant pathogens able to be transmitted by blood transfusions is widely implemented; however the presence of a series of pathogens in blood bank donations remained understudied. This is the case of some bacteria such as Leptospira spp. or Bartonella spp. Bartonella species are bloodborne, re-emerging organisms, capable of causing prolonged infections in animals and humans. Meanwhile, Leptospirosis is recognised as an emerging public health problem worldwide. Both infections are considered neglected tropical diseases.

Bartonella spp. are emerging bacterial pathogens which continue to achieve an ever-increasing level of clinical importance and awareness.  As Bartonella infections are common zoonoses in the U.S., the existence of Bartonella infection in Sweden, which had not been documented prior to the onset of our investigations, was of considerable interest. Initially, serological parameters of Bartonella infection were evaluated using a series of immunoblot analyses in an effort to elucidate the human humoral immunity induced by this pathogen.  An immunodominant epitope of B. henselae was identified, designated Bh83, which was hypothesized to be a conserved antigen during Bartonella infection. Further, an in-depth characterization of the immunoglobulin (Ig) isotype and IgG subclass response revealed a primary role of IgA and IgG1 in the immune induction during Bartonella infection. As the presence and degree of exposure to Bartonella spp. in Sweden was previously unknown, several studies were initiated to evaluate the level of seroreactivity to Bartonella spp. antigen in various subgroups of the Swedish human and animal population using indirect fluorescent-antibody assay (IFA).  A preliminary study of a select group of patients and blood donors represented the first documentation of Bartonella exposure in Sweden, as well as identified the first two cases of B. quintana-like infections in Sweden.  To assess the endemic level of antibodies to Bartonella spp. in Sweden, a large sampling of blood donor sera collected nationwide were subsequently analyzed for reactivity to six Bartonella strains.  The largest seroprevalence was to B. elizabethae, with relatively low baseline levels of reactivity to the other Bartonella antigens tested.  Correlated risk factors for infection included working outdoors and contact with cats, among others.  Further studies of sera obtained from elite orienteers and intravenous drug users in Sweden revealed a high level of exposure to Bartonella spp. in such cohorts, predominantly to B. elizabethae antigen.  Consistent with the aforementioned findings of elevated titers to B. elizabethae in humans, a high seroprevalence was found in cats, a known reservoir for Bartonella spp., as well. With relatively little known of the pathogenicity and epidemiology of Bartonella infection, these studies provide insight into various aspects of the distribution, risk factors for infection, specific humoral immune response to infection, and the prevalence of Bartonella spp. in human and cat populations in Sweden and serve as a framework for future studies.

La Enfermedad de Carrión, es causada por la bacteria Bartonella bacilliformis, que se transmite por varias especies del insecto vector del género Lutzomyia. No se conoce ningún reservorio animal, aparentemente el hombre es el único. Es una enfermedad endémica en el Perú y actualmente es considerada como reemergente debido a que su área geográfica se viene expandiendo. Es importante estudiar marcadores moleculares que sirvan para el diseño de métodos de diagnóstico rápido y efectivos, así como evaluar los antibióticos recomendados como medidas terapéuticas contra esta enfermedad, e indagar más acerca de las proteínas antigénicas de esta bacteria a fin de seleccionar antígenos candidatos para desarrollar, una vacuna contra esta enfermedad o pruebas de diagnóstico para su detección de laboratorio de mayor sensibilidad y especificidad que las actuales. IalB y FlaA son proteínas interesantes debido a su participación en la invasión bacteriana al eritrocito humano. / Carrion’s Disease is caused by Bartonella bacilliformis bacterium, and is transmitted by several species of the vector insect of the Lutzomyia genus. Animal reservoir is not known; seemingly the man is the only one. This disease is endemic in Peru and is actually considered as re-emergent because their geographical distribution comes expanding. Thus it is important to study its molecular markers for established diagnosis methods that are quick and effective. On the treatment aspect, it is necessary to evaluate the antibiotics used against the disease. On the therapeutic topic, more research is required regarding the antigenic proteins from this bacterium; this fact is important to develop future vaccine against this disease or diagnostic assay for their detection in the laboratory or to improve the sensitivity and specificity of the current methods. IalB and FlaA are interesting proteins due to than they are involved in the bacterial invasion to the human erythrocyte.

Bartonella bacilliformis es el agente causal de la enfermedad de Carrión. Actualmente, en el Perú los casos se siguen presentando y se ha reportado fallas en el tratamiento. Hay muy pocas investigaciones acerca de la susceptibilidad a antimicrobianos, y no se tiene estandarizada una prueba de antibiograma para esta bacteria. Asímismo, aún no se conocen los mecanismos de resistencia ni las secuencias de los genes asociados a ellas en cepas nativas circulantes de Bartonella bacilliformis. Por ello, el objetivo fue caracterizar molecularmente los genes asociados a la resistencia antimicrobiana en cepas circulantes en zonas endémicas para evaluar si presentan o no mutaciones en estos genes. Se estandarizó una prueba de antibiograma utilizando la cepa B. bacilliformis CIP 57.18 colocándose los discos de antibióticos comerciales: ciprofloxacina (CIP, 5 µg), rifampicina (RD, 30 µg), eritromicina (E, 15 µg), gentamicina (CN, 15 µg), doxiciclina (DO, 30 µg), cloranfenicol (30 µg), ácido nalidíxico (AN, 5 µg) y levofloxacina (LVX, 5 µg). Todas las cepas ensayadas mostraron resistencia al ácido nalidíxico pero sensibilidad a ciprofloxacina, a excepción de la cepa USM-LMM-002 que presentó susceptibilidad disminuida a ciprofloxacina. El análisis de las proteínas que codifican los genes gyrA y parC implicados en la resistencia a estos antibióticos, determinó que ambos presentan una diferencia aminoacídica (Ser83 por Ala) en la región del QRDR en relación a la GyrA y ParC de E. coli. Interesantemente, esta Ala83 en GyrA y ParC sólo proporciona resistencia a ácido nalidíxico y no a ciprofloxacina, siendo necesaria la acumulación de mutaciones en gyrA para la resistencia a ciprofloxacina. Para los demás antibióticos, las cepas fueron sensibles y en el secuenciamiento de los genes (rpoB, L4 y 16S ribosomal) asociado a dichas resistencia no se encontró ninguna mutación. La resistencia constitutiva al ácido nalidíxico es una propiedad de B. bacilliformis, y sería un buen indicador de la susceptibilidad a las fluoroquinolonas pero su resistencia no está relacionada a la resistencia de otras fluoroquinolonas. Además, la sensibilidad a otros antibióticos como doxiciclina, rifampicina, cloranfenicol y gentamicina podría indicar alternativas en el tratamiento de esta enfermedad aunque ensayos clínicos serian necesarios. / Bartonella bacilliformis is the causative agent of Carrion's disease. Currently in Peru are still present cases and are reported failures in treatment. There is very little research on antimicrobial susceptibility, nor has been standardized a test of sensitivity to the bacteria. Also, are not yet known resistance mechanisms and the sequences of genes associated with them in circulating native strains of Bartonella bacilliformis. Therefore, the objective was molecularly characterize the genes associated with antimicrobial resistance in isolates circulating in endemic areas to assess whether present or not mutations in these genes. Susceptibility testing was standardized using the strain of B. bacilliformis CIP 57.18 placing commercial antibiotic discs: ciprofloxacin (CIP, 5 µg), rifampin (RD, 30 µg), erythromycin (E, 15 µg), gentamicin (CN, 15 µg), doxycycline (DO, 30 µg), chloramphenicol (30 µg), nalidixic acid (NA, 5 µg) and levofloxacin (LVX, 5 µg). All strains tested showed resistance to nalidixic acid but sensitive to ciprofloxacin, except for strain USM-LMM-002 that presented reduced susceptibility to ciprofloxacin. The analysis of the proteins encoded by gyrA and parC genes involved in resistance to these antibiotics, determined that both have an aminoacid difference (Ser83 by Ala) in the QRDR region in relation to the GyrA and ParC of E. coli. Interestingly, the Ala83 in GyrA and ParC only provides resistance to nalidixic acid and not to ciprofloxacin, being necessary the accumulation of mutations in gyrA for ciprofloxacin resistance. For other antibiotics, the strains were sensitive and sequencing of genes (rpoB, and 16S ribosomal L4) associated with such resistance, there was no mutation. Constitutive resistance to nalidixic acid is a property of B. bacilliformis, and would be a good indicator of susceptibility to fluoroquinolones but this resistance is not related to resistance to other fluoroquinolones. In addition, sensitivity to other antibiotics such as doxycycline, rifampicin, chloramphenicol and gentamicin might suggest alternatives in the treatment of this disease, although clinical trials are needed.

Presentación de Trabajo Final para el curso Informática para Ciencias de la Salud (ME50) 2010-1

Presentación de Trabajo Final para el curso Informática para Ciencias de la Salud (ME50) 2010-1

Diss. (sammanfattning) Uppsala : Uppsala universitet, 2007. / Härtill 5 uppsatser.