ABSTRACT. Histoplasmosis, one of the most frequent endemic mycoses in the Americas, is caused by the inhalation of airborne conidia of Histoplasma capsulatum. Better understanding of the distribution of this fungus in the environment is important for the development of appropriate public health measures to prevent human infections. Previously, we used Hc100 nested polymerase chain reaction (PCR) to identify H. capsulatum DNA in 10% of environmental samples in Colombia. Here, we validate a 100-kDa real-time PCR assay for the detection of this fungus in the environment. Using this method, we identified H. capsulatum DNA in 80% of samples of raw organic materials, such as chicken manure, soil from caves, and bird and bat guano, as well as in 62% of samples of organic fertilizer that underwent the composting process. We demonstrated that 100-KDa real-time PCR is a useful tool for environmental surveillance that can be used to identify the potential reservoirs of H. capsulatum and to prevent outbreaks, especially in people with the higher risk of exposure, such as spelunkers, farmers, poultry manure collectors, and anyone who handle organic fertilizers or bat and bird excreta.

Minimal Residual Disease Assessment and Allogeneic Transplantation as Consolidation Therapy in Acute Lymphoblastic Leukemia Abstract Introduction: Detectable minimal residual disease (MRD) after therapy for acute lymphoblastic leukemia (ALL) is the strongest predictor of hematologic relapse (1-4). Colombian reports of adults with ALL showed poor survival rates, the median OS was 12 months and the median PFS was only 7 months (5,6). Most reports from Colombia as well as Latin America did not include patients that had a transplant as an initial consolidation strategy and used MRD as prognostic factor. (7-9). The objective of the study was to assess progression free survival (PFS) and overall survival (OS) of patients with ALL according with MRD status at end of induction therapy. Methods: A retrospective cohort comparing PFS and OS in adults with de novo ALL, according to the MRD status at the end of induction chemotherapy, and the type of post-induction consolidation strategy used. This research work had the endorsement of the ethics and research committees of the institutions where it was carried out. Results: Were included 165 adults with ALL in the MRD investigational group, the basal characteristics of the population are described in table 1.There were 73 patients in the MDR negative group and 92 in the MDR positive group. Median PFS for the MRD positive group was 11 months (CI 95% 11.7 - 22.2), and not reached for the MRD negative group (p<0.001). At three-years PFS was 18% and 55 %, respectively (p<0,001). Fig 1. The median OS for MRD positive patients was 16 months (CI 95%, 8.8 - 23.15) and was not reached in the MRD negative group. At three-years, OS was 26 % and 51 % for the former and latter group, respectively. Fig 2. Among subjects that were not transplanted, median PFS was 21 months for MRD negative and 9 months MRD positive patients (p<0.001). Fig 3A. Median of PFS was not reached in either group, whereas 3-year PFS was 64% for MRD negative and 70% for MRD positive patients when transplantation in first remission was used (p=0.861). Fig 3B. For the patients with only chemotherapy for consolidation treatment, the median OS were 16 months (IC 95% 11.32 to 20.67) for MRD positive, and 27 months IC 95% (20.91 to 33.09) in negative MRD. (p=0.004). The 3-year OS 34% for MRD negative and 22% for MRD positive patients. Fig 4A. The individuals with allogeneic transplantation during first remission overcome de risk of death in the positive MRD status. The median OS were 16 months for positive MRD and transplantation and not reached in negative MRD group, The 3 year OS were 74% for MRD negative and 49% for MRD positive patients. (p=0.187). Fig 4B. Multivariate analysis At multivariate analysis, the Negative MRD status and transplantation in first remission were associated with better PFS and OS, in these patients with ALL de novo, Table 2. Conclusion: MRD status at end of induction is an independent prognostic factor for PFS and OS in adult ALL. Allogeneic transplantation in first remission may overcome the adverse prognostic impact MRD. This is the first report that verifies the impact of MRD, on progression free survival and overall survival in the Colombian population. Disclosures No relevant conflicts of interest to declare.

The thermally dimorphic fungus Histoplasma capsulatum is the causative agent of histoplasmosis, one of the most prevalent endemic mycosis in the Americas. In tropical regions, agro-ecosystems require organic matter replacement, therefore, the use of organic fertilizers has increased disregarding the fact that certain number of such fertilizers might be contaminated with the fungus, and with their handling resulting in human cases and even outbreaks of histoplasmosis. Additionally, in Colombia, chicken manure is the most common raw material used in the production of organic fertilizers. In this work, we reported the isolation of this fungus from chicken manure, and genetically compared with 42 clinical isolates. The genetically compared environmental isolates grouped together with the clinical ones. Our result suggests that chicken manure may be one of H. capsulatum infection sources. Also, the phylogenetic analyses done with other H. capsulatum isolates indicate that the Colombian isolates are widely distributed in the relational tree thus reveling towards the great genetic diversity among the H. capsulatum Colombian isolates.

Sporothrix schenckii is a thermodimorphic fungal pathogen with a high genetic diversity. In this work, we present the assembly and similarity analysis of the whole-genome sequences of two clinical isolates from Colombia of S. schenckii sensu stricto.

The thermally dimorphic fungus Histoplasma capsulatum is the causative agent of histoplasmosis, which is the most prevalent endemic mycosis in America. The replacement of organic matter in agro-ecosystems is necessary in the tropics, and the use of organic fertilizers has increased. Cases and outbreaks due to the presence of the fungus in these components have been reported. In Colombia, chicken manure is the most common raw material in the organic fertilizers production. In this work, we reached the isolation of the fungus from a chicken manure. Then, we were able to compare genetically 3 environmental isolates with 42 Colombian human clinical isolates. The genetic comparison showed the environmental isolates grouping together with the clinical isolates. This result suggests chicken manure as one of the infection source with H. capsulatum . Also, the phylogenetic analysis using another H. capsulatum isolates from databases showed that the Colombian isolates widely distributed in the relation tree. This result pointed out the great genetic diversity among H. capsulatum Colombian population.

Histoplasma capsulatum is the causative agent of histoplasmosis and this fungus inhabits soils rich in phosphorus and nitrogen that are enriched with bird and bat manure. The replacement of organic matter in agroecosystems is necessary in the tropics, and the use of organic fertilizers has increased. Cases and outbreaks due to the presence of the fungus in these components have been reported. The Instituto Colombiano Agropecuario resolution 150 of 2003 contains the parameters set by the Colombian Technical Standard (NTC 5167) on the physicochemical and microbiological features of fertilizers, but it does not regulate the search for H. capsulatum. The aim of this study was to demonstrate H. capsulatum presence in organic fertilizers by nested polymerase chain reaction (PCR). A total of 239 samples were collected: 201 (84.1%) corresponded to organic fertilizers, 30 (12.5%) to bird excrement, and 8 (3.4%) to cave soils. The Hc100 nested PCR had a detection limit of 0.1 pg/µL and a specificity of 100%. A total of 25 (10.5%) samples were positive and validated by sequencing. Seven of the positive samples represented locations where H. capsulatum was previously detected, suggesting the persistence of the fungus. No significant correlations were detected between the physicochemical and microbiological parameters with the presence of H. capsulatum by nested PCR, indicating the fungus existence in organic fertilizers that complied with the NTC 5167. The Hc100 nested PCR targeting H. capsulatum standardized in this work will improve the evaluation of organic fertilizers and ensure the prevention of outbreaks and cases due to manufacturing, marketing, and use of fertilizers contaminated with H. capsulatum.

Introduction: In order to carry out an autologous transplantation, hematopoietic stem cells should be mobilized to peripheral blood and later collected by apheresis. The CD34+ cell count is a tool to establish the optimal time to begin the apheresis procedure. Objective: To evaluate the association between peripheral blood CD34+ cell count and the successful collection of hematopoietic stem cells. Materials and methods: A predictive test evaluation study was carried out to establish the usefulness of peripheral blood CD34+ cell count as a predictor of successful stem cell collection in patients that will receive an autologous transplantation. Results: 77 patients were included (median age: 49 years; range: 5-66). The predominant baseline diagnosis was lymphoma (53.2 %). The percentage of patients with successful harvest of hematopoietic stem cells was proportional to the number of CD34+ cells in peripheral blood at the end of the mobilization procedure. We propose that more than 15 CD34+ cells/μL must be present in order to achieve an adequate collection of hematopoietic stem cells. Conclusion: Peripheral blood CD34+ cell count is a useful tool to predict the successful collection of hematopoietic stem cells.

Introduction. Histoplasmosis, a fungal disorder characterized by a wide spectrum of manifestations that range from subclinical infections to disseminated processes, affects both immunocompetent and immunosuppressed individuals. Histoplasmosis is not a reportable disease in Colombia and consequently, a survey was designed to collect histoplasmosis cases diagnosed in the country.Objective. The aim of this work was to analyze the data collected from 1992 to 2008.Materials and methods. The survey included demographic data, risk factors, clinical manifestations, imaging data, diagnostic methods and antifungal treatment. Patients were grouped according to risk factors and comparisons of the various findings were done.Results. A total of 434 surveys were gathered from 20 of the country's Departments. Most patients (96.1%) were adults, 77% were males with a mean age of 38.4 years. Only 3.9% were children less than 15 years of age. In the adult population, AIDS was reported in 70.5% of the cases; additionally, in 7.0% patients other immunosuppressive conditions were informed. The most frequent clinical manifestations were fever (76.1%), cough (54.8%) and constitutional symptoms (56.8%). X rays abnormalities were represented mainly by infiltrates (65.9%) and nodules (17.1%). Diagnosis was made by microscopic observation of H. capsulatum in 49.6% patients, by culture in 58.0% and by serological test in 14.6% cases. Antifungal use was recorded in 52.5% cases.Conclusions. Histoplasmosis is frequent in Colombia, especially in certain risk factor groups such as the HIV-infected population. Data collected from this large number of cases has allowed valid comparisons on various aspects of histoplasmosis in Colombia.

Paracoccidioides brasiliensis has been classified in the phylum Ascomycota, order Onygenales, family Ajellomycetaceae, even in the absence of a known sexual cycle or mating system. The objective of this work was to determine the presence of the mating type locus in 71 P. brasiliensis isolates from various sources. A PCR assay using specific primers for the MAT 1 gene was developed and applied for the detection of such genes. Two heterothallic groups (MAT1-1 or MAT1-2) were recognized and, in some isolates, gene expression was confirmed indicating the existence of a basal gene expression. The distribution of two mating type loci in the studied population suggested that sexual reproduction might occur in P. brasiliensis. This finding points towards the possibility of applying a more precise definition of the concept of biological species to P. brasiliensis. Further studies should be conducted to confirm the sexual capacity of this fungus and its implications among phylogenetic species and geographical distribution.

Continuous real-time monitoring of the adequacy of cerebral perfusion can provide important therapeutic information in a variety of clinical settings. The current clinical availability of several non-invasive near-infrared spectroscopy (NIRS)-based cerebral oximetry devices represents a potentially important development for the detection of cerebral ischaemia. In addition, a number of preliminary studies have reported on the application of cerebral oximetry sensors to other tissue beds including splanchnic, renal, and spinal cord. This review provides a synopsis of the mode of operation, current limitations and confounders, clinical applications, and potential future uses of such NIRS devices.

Se determinó la sensibilidad al fluconazol y al voriconazol de aislamientos de Candida spp. obtenidos de la mucosa oral de 54 pacientes con sida hospitalizados en la ESE Hospital La María (Medellín, Colombia). Además, se comprobó la especie de tales aislamientos. Los pacientes eran todos adultos (promedio de 40,5 años, rango de 23 a 56) y la mayoría (77,8%) hombres. En 40 (71,1%) de ellos se obtuvo crecimiento de Candida spp. y en 6 (11,1%) se aisló más de una especie de Candida. La clasificación a especie reveló C. albicans, C. krusei, C. tropicalis, C. parapsilosis y C. glabrata. La determinación de la sensibilidad in vitro se hizo por difusión en agar con las especificaciones del CLSI de Estados Unidos (M44P). El 72,9% de los aislamientos de Candida spp. fueron sensibles al fluconazol; 6,3%, sensibles dependientes de la dosis, y 20,8%, resistentes. Para el voriconazol, 89,6% fueron sensibles; 8,3%, sensibles dependientes de la dosis y 2,1%, resistentes. La resistencia se observó en C. albicans y C. krusei. Estos resultados demuestran la efectividad de ambos antimicóticos y fue mayor la correspondiente al voriconazol. Igualmente se señala la importancia de las pruebas in vitro puesto que en pacientes con VIH/sida pueden encontrarse especies resistentes. Se ratifica la importancia de la clasificación por especie de las levaduras del género Candida.Abstracts The susceptibilities to Fluconazol (FCZ) and voriconazol (VCZ) susceptibility of Candida spp. isolates, obtained from the oral mucosa of 54 AIDS patients hospitalized in Hospital La María (Medellín, Colombia) were assesed. Additionally, the yeast species were identified to species. Patients were all adults (mean, 40,5 years old, range 23 to 56) and most (77,8%) were males. Candida spp. was isolated from 40 of them (71.1 %) with 6 (11,1%) showing more than one species. The species identified were C. albicans., C. krusei, C. tropicalis, C. parapsilosis and C. glabrata. In vitro agar diffusion susceptibility testing was done according to the CLSI M44P specifications. Most of the Candida species (72.9%) were susceptible to fluconazole with 6,3% susceptible-dose-dependant (SDD) and 20.8% resistant. Voriconazole susceptibility was higher (89,6%) with 8,3% SDD and 2,1% resistant. Resistance was recorded for C. albicans and C. krusei isolates. The results demonstrate the effectiveness of both antifungals with voriconazole performing better. The importance of the in vitro susceptibility tests in HIV/aids patients harboring resistant species was demonstrated and indicated the need to classify all Candida isolate al Candida isolates to the species level.

We investigated angiogenesis, inflammatory cells accumulation and endogenous production of cytokines in sponge implants of tumor-bearing mice. Seven days after inoculation of Ehrlich tumor cells (2.5 × 106), sponge discs were implanted subcutaneously in the dorsa of mice to induce the formation of fibrovascular tissue. The implants of tumor-bearing and non tumor-bearing animals were assessed for neovascularization and leukocyte accumulation, together with levels of relevant cytokines, vascular endothelial growth factor VEGF), tumor necrosis factor alpha (TNF-α), CXCL1-3/KC and CCL2/JE. In the implants of tumor-bearing animals angiogenesis (assessed by hemoglobin content and VEGF levels in the implants) and leukocyte accumulation (assessed by myeloperoxidase -MPO- and N- acetylglucosaminidase-NAG-enzyme activities) were all significantly less than those in the implants of non tumor-bearing animals. Although the chemokine CXCL1-3/KC was lower in the implants of tumor-bearing animals, the chemokine CCL2/JE was increased in this group. The production of TNF-α in the implants was not modified by the presence of the subcutaneous tumor. The combination of the methodologies used in this study has provided a novel approach to investigate the interaction between two distinct proliferating tissues that share common features (angiogenesis, cell recruitment, inflammation) and has shown that the predominant inhibitory effect of a tumor mass over repair process is associated with altered cytokine production.

Acute and late convalescent sera (collected at day 5 of disease onset and 1 year later) from dengue fever (DF) and dengue hemorrhagic fever/dengue shock syndrome (DHF/DSS) laboratory confirmed cases, were tested for antibody-dependent cell-mediated cytotoxicity (ADCC) activity using dengue 1 (DENV-1) or dengue 2 (DENV-2) infected cells as target. All patients experienced their first dengue virus (DENV) infection 20 years before. ADCC activity was detected in acute sera from DHF/DSS but not in sera from DF patients. However, 1 year after illness, ADCC activity was observed in all cases. This preliminary report represents one of the few studies of ADCC in dengue patients and suggests that ADCC could be implicated in dengue pathogenesis.

1Department of Anesthesia, University of Western Ontario, Canada, 2Fundación Médica Santa Fe Bogotá, Colombia, 3Fundación Valle de Lili Cali, Colombia

We studied 52 patients with disseminated histoplasmosis, 30 with the acquired immunodeficiency syndrome (AIDS) (cohort 1) and 22 not co-infected with the human immunodeficiency virus (cohort 2). Demographic, clinical, laboratory, mycologic findings, as well as antifungal therapy and highly active antiretroviral (HAART), were analyzed. Skin lesions were significantly higher in cohort 1 than in cohort 2 (P = 0.001). Anemia, leukopenia, and an elevated erythrocyte sedimentation rate were also more pronounced in cohort 1 than in cohort 2 (P < 0.001). Histoplasma capsulatum was isolated more often in cohort 1 than in cohort 2 (P < 0.05) patients, but antibodies to H. capsulatum were detected more frequently in cohort 2 than in cohort 1 (P < 0.05). Itraconazole treatment was less effective in cohort 1 than in cohort 2 (P = 0.012). In cohort 1 patients, HAART improved response to antifungals when compared with individuals not given HAART (P = 0.003), who exhibited higher mortality rates (P = 0.025). Cohort 1 patients who were given dual antifungal and anti-retroviral therapies responded as well as the non-HIV patients in cohort 2, who were treated only with itraconazole. These results indicate the need to promote restoration of the immune system in patients with AIDS and histoplasmosis.