Search

Resumen La demencia con cuerpos de Lewy (DCL) es la segunda demencia degenerativa mas frecuente. Se caracteriza clinicamente por deficits de atencion y funciones visuoespaciales, fluctuaciones, alucinaciones visuales, parkinsonismo y alteraciones de la conducta durante el sueno REM. En la enfermedad de Parkinson avanzada, es muy frecuente la demencia asociada, que muchas veces es clinicamente similar a la DCL, con la que comparte alteraciones anatomopatologicas, por lo que se plantea que sean solo diferentes expresiones clinicas de una misma enfermedad. La demencia frontotemporal es la demencia degenerativa mas frecuente en personas jovenes. Puede comenzar con trastornos conductuales o en forma de afasias primarias. La variante conductual es la mas frecuente y se caracteriza por desinhibicion, apatia, perdida de empatia, comportamientos compulsivos e hiperoralidad. Las pruebas neuropsicologicas y las de neuroimagen pueden ser normales en fases iniciales. Las demencias vasculares constituyen el segundo tipo de demencia mas frecuente tras la enfermedad de Alzheimer. La alteracion precoz de las funciones ejecutivas, la historia de ictus, la presencia de factores de riesgo vascular y la progresion escalonada deben hacer sospechar esta entidad. La presencia de ictus y/o lesiones isquemicas de sustancia blanca apoyan el diagnostico.

Resumen Los trastornos del movimiento cursan frecuentemente con alteraciones cognitivas y neuropsiquiatricas asociadas. El primer paso consiste en averiguar si hay farmacos que pueden provocarlos y proceder a su retirada. La enfermedad neurodegenerativa que cursa con trastornos hipercineticos, deterioro cognitivo y alteraciones conductuales mas importante es la enfermedad de Huntington. El parkinsonismo secundario (farmacologico o de otra causa –hipotiroidismo, hidrocefalia, vascular, etc.—) requiere un tratamiento etiologico cuando sea posible. El adecuado diagnostico de los parkinsonismos neurodegenerativos tiene implicaciones terapeuticas y pronosticas importantes.

Mycotoxins may be present in nuts, coffee, cereals, and grapes, among other products. Increasing concerns about human health and environmental protection have driven the application of biological control techniques that can inhibit fungal contaminants. In this study, the growth inhibition of the ochratoxigenic fungus Aspergillus carbonarius Ac 162 was evaluated using 5 lactic acid bacteria (LAB). The LAB studied were Lactobacillus plantarum MZ801739 (J), Lactobacillus plantarum MZ809351 (31) and Lactobacillus plantarum MZ809350 (34), isolated in the Ivory Coast, and Lactobacillus plantarum MN982928 (3) and Leuconostoc citreum MZ801735 (23), isolated in Mexico. J, 31, 34, 3 and 23 are the internal strain codes from our laboratory. LAB were cultivated in De Man, Rogosa and Sharpe (MRS) broth, and different polyols (glycerol, mannitol, sorbitol, and xylitol) were added to the culture broth to stimulate the production of antifungal compounds. The fungal inhibition studies were performed using the poisoned food technique. The highest inhibition of A. carbonarius growth was obtained by cultivating L. plantarum MZ809351 in the presence of xylitol and glycerol. Under these conditions, 1 L of the L. plantarum MZ809351 cultures were used to identify antifungal compounds. The compounds were concentrated by solid-phase extraction and then characterized by GC–MS. In addition to 9-octadecenoic acid, 3 diketopiperazines or cyclic dipeptides were identified, including cyclo (Leu-Leu), cyclo (Pro-Gly) and cyclo (Val-Phe), which were compounds related to microbial antifungal activities. Xylitol and glycerol induced the production of these antifungal compounds against A. carbonarius Ac 162. On the other hand, adding xylitol and glycerol to the MRS broth reduced the Ochratoxin A (OTA) content to 56.8 and 54.7%, respectively. This study shows the potential for using L. plantarum MZ809351 as a biocontrol agent to prevent the growth of A. carbonarius and reduce the production of OTA in foods.

Affinity characterization is essential to develop reliable aptamers for tumor biomarker detection. For alpha-fetoprotein (AFP), a biomarker of hepatocellular carcinoma (HCC), two DNA aptamers were described with very different affinity. In this work, we estimate the dissociation constant of both of them by means of a direct assay on magnetic beads modified with AFP and electrochemical detection on carbon screen-printed electrodes (SPCE). Unlike previous works, both aptamers showed similar dissociation constant (Kd) values, in the sub&micro, M range. In order to improve the performance of these aptamers, we proposed the isothermal amplification of the aptamers by both terminal deoxynucleotidyl transferase (TdT) and rolling circle amplification (RCA). Both DNA amplifications improved the sensitivity and also the apparent binding constants from 713 nM to 189 nM for the short aptamer and from 526 nM to 32 nM for the long aptamer. This improvement depends on the true affinity of the binding pair, which ultimately limits the analytical usefulness.

A perchloric acid-soluble protein (PSP), named here tv-psp1, was identified in Trichomonas vaginalis. It is expressed under normal culture conditions according to expressed sequence tag (EST) analysis. On the other hand, Tv-PSP1 protein was identified by mass spectrometry with a 40% of identity to human PSP (p14.1). Polyclonal antibodies against recombinant Tv-PSP1 (rTv-PSP1) recognized a single band at 13.5 kDa in total protein parasite extract by SDS-PAGE and a high molecular weight band analyzed by native PAGE. Structural analysis of Tv-PSP1, using dynamic light scattering, size exclusion chromatography, and circular dichroism spectroscopy, showed a trimeric structure stable at 7 M urea with 38% α-helix and 14% β-sheet in solution and a molecular weight of 40.5 kD. Tv-PSP1 models were used to perform dynamic simulations over 100 ns suggesting a stable homotrimeric structure. Tv-PSP1 was located in the nucleus, cytoplasm, and hydrogenosomes of T. vaginalis, and the in silico analysis by Search Tool for the Retrieval of Interacting Genes/Proteins (STRING) showed interactions with RNA binding proteins. The preliminary results of RNA degradation analysis with the recombinant Tv-PSP1 showed RNA partial deterioration suggesting a possible putative ribonuclease function.

Global change drivers are currently affecting semiarid ecosystems. Because these ecosystems differ from others in biotic and abiotic filters, cues for plant regeneration and management derived from elsewhere may not be applicable to semiarid ecosystems. We sought to determine the extent to which regional variation in regeneration prospects of a long-lived semiarid keystone shrub depends on anthropogenic habitat degradation, plant-animal interactions and climate determinants. We investigated the regeneration ability (via population size structure, juvenile density and juvenile/adult ratio), fruit set and seed dispersal of Ziziphus lotus in 25 localities spanning the range of its threatened habitats in Spain. We dissected the relative contribution of different regeneration determinants using multiple regression and structural equation modelling. Population regeneration was extremely poor, and size structures were biased towards large classes and low juvenile densities and juvenile/adult ratios. Poor regeneration was often coincident with seed dispersal collapse. However, the positive effect of seed dispersal on population regeneration disappeared after considering its relationship with habitat degradation. Protected areas did have juveniles. Together, these data suggest that habitat degradation directly impacts juvenile establishment. Our results provide insights into habitat and species management at the regional level. Z. lotus populations are currently driven by persistence-based dynamics through the longevity of the species. Nonetheless, collapsed seed dispersal, poor regeneration and the removal of adults from their habitats forecast extinction of Z. lotus in many remnants. The extreme longevity of Z. lotus provides opportunities for recovery of its populations and habitats through effective enforcement of regulations.

Acanthamoeba culbertsoni trophozoites, previously isolated from a human keratitis case with severe intraocular damage, were maintained in axenic culture. Co-incubation of amoebae with MDCK cell monolayers demonstrated an apparent preference of the amoebae to introduce themselves between the cells. The trophozoites appeared to cross the cell monolayer through the tight junctions, which resulted in decreased trans-epithelial resistance (TER) measurements. Unexpectedly, after co-incubation of amoebae with hamster corneas, we observed that the trophozoites were able to cross the different cell layers and reach the corneal stroma after only 12 h of interaction, in contrast to other Acanthamoeba species. These observations suggest that this A. culbertsoni isolate is particularly pathogenic. Further research with diverse methodologies needs to be performed to explain the unique behavior of this Acanthamoeba strain.

Free-living amoebae of the genus Acanthamoeba are protozoa ubiquitously found in nature. Some species of the genus are potentially pathogenic for humans provoking keratitis in healthy individuals, often in contact lens wearers and opportunistic infections such as pneumonitis, fatal granulomatous encephalitis and skin infections, particularly in immunocompromised individuals. The pathogenic mechanisms of these amoebae are poorly understood, however it had been suggested that contact dependent mechanisms are important during invasion, regardless of the epithelia type, since amoebae penetrate epithelia separating tight junction (TJ). This study was undertaken to determine whether Acanthamoeba sp. (T4) damages the barrier function of the TJ in MDCK epithelial monolayers. Actin cytoskeleton staining and electron microscopy analyses were performed; paracellular permeability and TJ sealing were evaluated by apicobasolateral diffusion of ruthenium red and transepithelial resistance (TER) measurements; immunofluorescence and Western blot assays were performed to locate and estimate expression of TJ protein claudins 2 (Cldn2) and 4 (Cldn4). The results show that Acanthamoeba sp. crosses the MDCK monolayer without altering the actin cytoskeleton or the morphology of the cells. When trophozoites or conditioned medium interact with the monolayer, paracellular diffusion of ruthenium red increases. After 6 h, the amoebae, but not their conditioned medium, increase the TER, and Cldn2 is removed from the TJ, and its overall content in the cells diminishes, while Cldn4 is targeted to the TJ without changing its expression level. In conclusion Acanthamoeba (T4) crosses MDCK monolayer without damaging the cells, increasing permeability and TER through Cldn2 degradation, and redirecting Cldn4 to TJ. These results strongly suggest that contact-dependent mechanisms are relevant during amoebae invasion.

Objective In this study we analyzed the effects of transcranial magnetic stimulation (TMS) on sleep and on the self-perceived quality of life in epileptic patients. Methods A total of 24 male patients diagnosed with focal epilepsy were included in the study. Pharmacological treatment with levetiracetam was standardized at 2 g daily. Before TMS onset, all-night polysomnographic recording (PSG) was performed, and the Quality of Life in Epilepsy Inventory (QOLIE-31) was administered. Thereafter, patients underwent low-frequency repetitive TMS (1000 pulses/1 Hz) daily for 10 days. After the end of the treatment, a second polysomnographic study was performed, and the QOLIE-31 questionnaire was administered again. Results TMS induced a significant increase in sleep efficiency and in total sleep time, along with a decrease in sleep latency and the number of awakenings. In addition, the number of interictal discharges during sleep decreased significantly. Concerning the QOLIE-31 scale values, the patients showed great improvement in the self-perceived quality of life. Conclusion The present results indicate that TMS may mediate therapeutic effects in the treatment of patients with focal epilepsy, and that TMS treatment is accompanied by improvement of sleep patterns as well as improvement in self-perceived quality of life. However, a study that includes a control group undergoing sham stimulation is needed to confirm these findings.

Acanthamoeba keratitis (AK) is a sight-threatening corneal infection. The early symptoms include redness, pain, photophobia and intense tearing. Chronic infection usually progresses to stromal inflammation, ring ulcers, corneal opacification and hypopyon. Here we document an AK case in a high myopic 38-year-old woman from Mexico City, with a history of wearing contact lenses while swimming. Corneal scrapes cultures were positive only for amoebae, consequently a treatment including netilmicin 0.3% and oral itraconazole 100 mg/12 h was prescribed. The infection was resolved after 8 months, leaving a slight leucoma outside the visual axis, with a visual acuity of 20/150. In the laboratory, the amoebic isolate was axenized in PYG medium, with an optimal growth at 30 °C, and was identified morphologically as Acanthamoeba polyphaga according to the taxonomic criteria of Page (1988) and placed in the T4 group by genotyping. The virulence of this strain (40%) was determined by intranasal inoculation of 1 × 106/20 μl trophozoites in BALB/c mice recovering from brain, proving their invasion ability and by the interaction with monolayers of epithelial cells of the established MDCK line of canine kidney origin (1:2 ratio of interaction), at 1, 3, 6, 8 and 24 h; trophozoites migrated to cell junctions inducing few lytic zones. In addition to the biological characterization, in vitro drug sensitivity tests were performed using chlorhexidine, itraconazole, netilmicin and voriconazole. Results revealed that voriconazole was the most effective compound. A. polyphaga remains as one of the most frequently isolated species producing AK. The treatment of AK case using netilmicin and oral itraconazole solved the disease, but the healing process was wide-ranging (8 months). The use of voriconazole and chlorhexidine may be an alternative treatment of future AK cases in Mexico.

The metabolic and kinetic behaviour of a nitrification process in the presence of 2-chlorophenol (2-CP) was evaluated in two sequencing batch reactors (SBR1, SBR2) inoculated with nitrifying sludge previously exposed to phenolic compounds. The SBR1 was inoculated with sludge previously exposed to 2-CP, while the SBR2 was inoculated with sludge previously exposed to

Additional morphological features of Balamuthia mandrillaris observed by light and electron microscopy are reported. Trophozoites were extremely pleomorphic: their cell shapes ranged from rounded to elongated and sometimes they appeared exceptionally stretched out and branched. By transmission electron microscopy it was possible to observe two different cytoplasmic areas, the ectoplasm and the endoplasm and often sections of rough endoplasmic reticulum were found in the transition zone. The cytoplasm was very fibrogranular and most of the organelles typically found in eukaryotic cells were observed. A particular finding was the presence of numerous mitochondria with a different structure from those of other free-living amoebae. The observations reported here may reinforce the morphological knowledge of this amoeba and provide a background for further analyses.

Uno de los aspectos más relevantes en Psicopatología es la clasificación de los trastornos mentales. La caracterización de los distintos problemas psicopatológicos en manuales diagnósticos estandarizados como los DSM publicados por la Asociación de Psiquiatría Americana (APA) o las Clasificaciones Internacionales de las Enfermedades (CIE) publicadas por la Organización Mundial de la Salud (OMS) tiene una influencia fundamental en aspectos relevantes en la psiquiatría y la psicología clínica como el diagnóstico, el pronóstico, el tratamiento o la investigación. La publicación del DSM-5 va a suponer uno de los hitos más importantes en el ámbito de la salud mental en los últimos años. La revisión del DSM-IV ha puesto en cuestión diferentes aspectos fundamentales de las clasificaciones psiquiátricas vigentes (p. ej., estructura de la clasificación, enfoque prototípico, cuestiones relacionadas con la etiología, etc.). Fruto de esta revisión se han producido importantes modificaciones en algunos de estos aspectos. El objetivo de este trabajo es llevar a cabo una revisión general de los cambios que implica la sustitución del DSM-IV por el nuevo DSM-5. Estos cambios han sido especialmente sustanciales en la clasificación de los trastornos de personalidad (TP). Por ello, a fin de ejemplificar las modificaciones sufridas por el DSM, nos centramos en este grupo de trastornos. Finalizamos el artículo con una reflexión acerca de las repercusiones de la publicación del DSM-5 One of the most important topics in Psychopatology is the classification of mental disorders. The description of different psychopathological problems in standardized diagnostic manuals such as the DSM, published by the American Psyquiatric Association (APA), or the icd, published by the World Health Organization (WHO), strongly affects central aspects related to psychology and psychiatry, such as diagnosis, prognosis, treatment and research. The next publication of DSM-5 is expected to become an important milestone in mental health. The review has questioned several major aspects of current psychiatric classifications (e.g. classification framework, prototypical approach, different questions about etiology, etc.). As a result, important changes in some of these aspects have been done. The aim of this paper is to carry out a general review of the modifications that involves the replacement of DSM-IV by the new DSM-5. These changes have been especially important in the classification of personality disorders. For this reason, as an example, we will focus on the changes undergone by personality disorders. As a conclusion, we end our article with a reflection about the implications of the publication of DSM-5.

La fibromialgia (FM) y otros síndromes de dolor crónico constituyen un importante problema de salud por el deterioro de la actividad y la calidad de vida que producen. Estos problemas requieren un abordaje multidisciplinar, sin embargo, el tratamiento actual todavía es insuficiente. Una forma de mejorar las intervenciones podría ser el uso de las tecnologías de la información y la comunicación (TIC). El grupo de investigación Labpsitec ha creado TEO (Terapia Emocional On Line), un sistema abierto y autoaplicado que permite recoger datos del paciente en tiempo real en su ambiente natural, crear sesiones de tratamiento personalizadas y enviarlas a los pacientes mediante Internet. Con esta herramienta pretendemos trabajar el mantenimiento de las estrategias aprendidas por los pacientes ya que, en ocasiones, dejan de practicarlas a medio o largo plazo una vez finalizados los tratamientos. El objetivo de este trabajo es describir el programa TEO-FM y presentar datos preliminares de la aceptación del mismo por parte de tres pacientes (edad media de 57,33 años [dt = 0,577]), con diagnóstico de síndromes de dolor crónico. Las tres utilizaron TEO de forma autoaplicada durante 4 semanas tras finalizar un tratamiento presencial. Todas manifestaron un alto grado de satisfacción y una opinión muy positiva de la herramienta. Estos resultados preliminares ayudan a continuar con la validación de TEO como una herramienta para que los pacientes consoliden lo aprendido en las sesiones, contribuyendo a su calidad de vida y su bienestar. Fibromyalgia (FM) and other chronic pain syndromes are a major health problem due to the decline in activity and quality of life they produce. These problems require a multidisciplinary approach; however, the current interventions present still limitation that could be improved. A way to progress in this line is to include the support of Information and Communication Technologies (ICT). The research group Labpsitec has created TEO (Emotional Therapy On-Line), an open and self-applied system that allows collecting patient data in real time in their natural environment, to create custom treatment sessions and to send them to patients via Internet. With this tool we try to obtain the maintenance of the strategies learned by patients because sometimes they stop practicing them at mid-and long-term once the treatment is over. The aim of this paper is to describe the program and present data of TEO-FM preliminary acceptance by three patients (mean age 57.33 years [SD = 0.577]), with a diagnosis of chronic pain syndromes. These three patients used TEO in a self administered mode for 4 weeks after finishing a face-to-face treatment program. All reported a high degree of satisfaction and a very positive opinion about the tool. These preliminary results help to continue the validation of TEO as a tool for patients to consolidate what has been learned in the sessions, contributing to their quality of life and welfare.

El dolor lumbar crónico afecta a un gran número de personas y constituye un grave problema de salud pública pues en muchos casos no están del todo claros ni las causas ni el adecuado tratamiento para estos pacientes. Asociado a este problema, se encuentran diversos factores psicológicos que hacen que el problema se agrave. Por otra parte, es habitual encontrar problemas en el mantenimiento de las estrategias aprendidas en terapia y por ello, el grupo Labpsitec ha creado la plataforma POET que permite seguir trabajando los contenidos aprendidos desde casa una vez terminadas las sesiones de terapia. El objetivo de este trabajo es describir el programa POET y presentar datos preliminares de la aceptación por parte de uno de los primeros pacientes con dolor lumbar crónico que lo han utilizado, cuya valoración del sistema ha sido muy positiva tanto en satisfacción como en utilidad. Estos datos nos ayudan a seguir con la validación de esta herramienta para que los pacientes continúen en contacto con lo aprendido en terapia y puedan ponerlo en práctica en su día a día con el fin de mejorar su calidad de vida. Chronic back pain affects a large number of people and constitutes a serious public health problem because in a lot of cases the causes and the appropriate treatment for these patients are not clear. Related with this, there are several psychological factors that make the problem worse. Moreover, it is common to find problems in maintaining the strategies learned in therapy and therefore, the group Labpsitec has created POET, a system that allows working from home the contents learned in the therapy sessions. The aim of this study is to describe POET and show preliminary data from the acceptance of one of the first patients with chronic low back pain who used it. The patient reported a positive opinion about the system. This data helps us to continue with the validation of this tool in order to provide patients with a system that could help them to maintain the treatment outcomes in their daily lives with the final aim of improving their quality of life.

Introduction: Fibromyalgia (FM) is a chronic musculoskeletal pain condition of unknown etiology, often accompanied by fatigue, sleep disturbance and depressed mood. It is a complex syndrome involving biological, psychological and social factors which causes a negative impact in the patient´s quality of life. Biopsychosocial models of pain hypothesize that patient attitudes and beliefs about pain play a key role in the adjustment to chronic pain. For that reason, a number of self-report instruments have been developed to assess those constructs. The Survey of Pain Attitudes (SOPA) is one of the most commonly used measures of pain beliefs having shown good psychometric properties. For this study, we used the SOPA-B (brief version) to preliminary test its validity in a sample of Spanish women diagnosed with FM. Aim: To develop a Spanish adaptation of the SOPA-B and examine its factor structure preliminarily Methods: 258 female patients with FM (ACR, 1990) were administered the SOPA-B. Results: Factor analyses supported a six-factor structure: Solicitude, Emotion, Disability, Harm, Control and Medical Procedures, consisting of 28 items. The Spanish-SOPA-B showed to be a reliable measure as demonstrated by the scales Cronbach’s alpha (ranging from 0.83 to 0.60). Conclusions: The adaptation and translation process of the SOPA, led to the confirmation of five of the original scales of this questionnaire in a different cultural group. These findings are promising and indicate that the Spanish-SOPA-B has good reliability and validity properties. Further studies are needed to confirm these preliminary findings Introducción: La fibromialgia (FM) es una condición de dolor musculoesquelético crónico de etiología desconocida, a menudo acompañado de fatiga, alteraciones del sueño y estado de ánimo depresivo. Es un síndrome complejo que involucra factores biológicos, psicológicos y sociales que provoca un impacto negativo en la calidad de vida del paciente. Modelos biopsicosociales del dolor tienen la hipótesis de que las actitudes del paciente y sus creencias acerca del dolor juegan un papel clave en la adaptación al dolor crónico. Por esa razón, una serie de instrumentos de autoinforme se han desarrollado para evaluar aquellos constructos. El Cuestionario de Actitudes hacia el Dolor (SOPA) es una de las medidas más utilizadas para evaluar las creencias hacia el dolor y ha demostrado tener buenas propiedades psicométricas. Para este estudio, se utilizó la versión breve del SOPA-B para evaluar su validez en una muestra de mujeres españolas con diagnóstico de FM. Objetivo: Desarrollar una adaptación española del cuestionario SOPA-B y examinar su estructura factorial de manera preliminar. Método: A 258 mujeres con FM (ACR, 1990) se les administró el cuestionario SOPA-B. Resultados: Los análisis factoriales dieron como resultado una estructura de seis factores: Solicitud, Emoción, Discapacidad, Daño, Control y Procedimientos Médicos, que consta de 28 ítems. La versión española del SOPA-B demostró ser una medida fiable como lo demuestran las escalasalfa de Cronbach (que van desde 0,83 hasta 0,60). Conclusiones: El proceso de adaptación y traducción del cuestionario SOPA confirmó cinco de las escalas originales de este cuestionario en un grupo cultural diferente. Estos hallazgos son prometedores e indican que la versión en español del SOPA-B es un instrumento fiable y válido. Se necesitan más estudios para confirmar estos hallazgos preliminares.

Granulomatous amoebic encephalitis (GAE) is a chronic, difficult to resolve infection caused by amphizoic amoebae of the genus Acanthamoeba , which in most cases occurs in immunosuppressed persons or with chronic diseases such as diabetes. In this study, we describe the early events of A. culbertsoni infection of GAE in diabetic mice model. Diabetes was induced in male BALB/c mice, with a dose of streptozotocin (130 mg/kg). Healthy and diabetic mice were inoculated via intranasal with 1 × 10 6 trophozoites of A. culbertsoni . Then were sacrificed and fixed by perfusion at 24, 48, 72 and 96 h post-inoculation, the brains and nasopharyngeal meatus were processed to immunohistochemical analysis. Invasion of trophozoites in diabetic mice was significantly greater with respect to inoculated healthy mice. Trophozoites and scarce cysts were immunolocalized in respiratory epithelial adjacent bone tissue, olfactory nerve packets, Schwann cells and the epineurium base since early 24 h post-inoculation. After 48 h, trophozoites were observed in the respiratory epithelium, white matter of the brain, subcortical central cortex and nasopharyngeal associated lymphoid tissue (NALT). At 72 h, cysts and trophozoites were immunolocalized in the olfactory bulb with the presence of a low inflammatory infiltrate characterized by polymorphonuclear cells. Scarce amoebae were observed in the granular layer of the cerebellum without evidence of inflammation or tissue damage. No amoebas were observed at 96 h after inoculation, suggesting penetration to other tissues at this time. In line with this, no inflammatory infiltrate was observed in the surrounding tissues where the amoebae were immunolocalized, which could contribute to the rapid spread of infection, particularly in diabetic mice. All data suggest that trophozoites invade the tissues by separating the superficial cells, penetrating between the junctions without causing cytolytic effect in the adjacent cells and subsequently reaching the CNS, importantly, diabetes increases the susceptibility to amoebae infection, which could favor the GAE development.

Entamoeba histolyticais the causative agent of human intestinal and liver amebiasis. The extraordinary phagocytic activity ofE. histolyticatrophozoites has been accepted as one of the virulence mechanisms responsible for their invasive capacity. The recognition of the noninvasiveEntamoeba disparas a different species has raised the question as to whether the lack of pathogenic potential of this ameba correlates with a limited phagocytic capacity. We have therefore compared the process of erythrophagocytosis in both species by means of light and video microscopy, hemoglobin measurement, and the estimation of reactive oxygen species (ROS). In the present study, we confirmed thatE. disparhas lower erythrophagocytic capacity. We also observed by video microscopy a new event of erythrocyte opsonization-like in both species, being more characteristic inE. histolytica. Moreover,E. disparshowed a lower capacity to produce ROS compared with the invasive species and also showed a large population of amoebae that did not engulf any erythrocyte over time. Our results demonstrate thatE. histolyticahas a higher phagocytic capacity thanE. dispar, including a higher rate of production of ROS in the course of ingesting red blood cells.

Observations on cultured Acanthamoeba royreba trophozoites and in vitro cytopathogenicity of this amoeba are described. In culture, amoebae were active, pleomorphic and moved on the substrate by producing endocytic structures and emitting slight cytoplasmic microprojections from the cell surface. These projections were formed by hyaline cytoplasm and they were related to motion structures such as acanthopodia and lamellipodia, in which actin provides a framework that allows rapid changes in morphology. In the cytoplasm abundant vacuoles of different size and content were seen. By means of electron microscopy, it was possible to observe the compact fibrogranular appearance of the cytoplasm, along with the main cellular organelles such as the Golgi complex, the endoplasmic reticulum, digestive vacuoles, mitochondria and contractile vacuoles. Incubation of MDCK epithelial cell monolayers with conditioned medium did not produce a significant structural damage to the monolayer, even after 24h of incubation. When the trophozoites were incubated with the target cells the monolayer exhibited a clear injury created by the amoebae, which produced focal damage. Nevertheless, the rest of the monolayer appeared to remain intact, suggesting that a contact-dependent interaction is necessary to damage the target cells. These observations demonstrate the low invasive capacity of this amoeba.

BackgroundShiga toxin-producingEscherichia coli(STEC) is one of the most common and widely distributed foodborne pathogens that has been frequently implicated in gastrointestinal and urinary tract infections. Moreover, high rates of multiple antibiotic-resistantE. colistrains have been reported worldwide. Due to the emergence of antibiotic-resistant strains, bacteriophages are considered an attractive alternative to biocontrol pathogenic bacteria. Characterization is a preliminary step towards designing a phage for biocontrol.MethodsIn this study, we describe the characterization of a bacteriophage designated phiC119, which can infect and lyse several multidrug-resistant STEC strains and someSalmonellastrains. The phage genome was screened to detect thestx-genes using PCR, morphological analysis, host range was determined, and genome sequencing were carried out, as well as an analysis of the cohesive ends and identification of the type of genetic material through enzymatic digestion of the genome.ResultsAnalysis of the bacteriophage particles by transmission electron microscopy showed that it had an icosahedral head and a long tail, characteristic of the familySiphoviridae. The phage exhibits broad host range against multidrug-resistant and highly virulentE. coliisolates. One-step growth experiments revealed that the phiC119 phage presented a large burst size (210 PFU/cell) and a latent period of 20 min. Based on genomic analysis, the phage contains a linear double-stranded DNA genome with a size of 47,319 bp. The phage encodes 75 putative proteins, but lysogeny and virulence genes were not found in the phiC119 genome.ConclusionThese results suggest that phage phiC119 may be a good biological control agent. However, further studies are required to ensure its control of STEC and to confirm the safety of phage use.

Introducción: En los últimos años ha habido un interés creciente en abordar el trata-miento de los trastornos emocionales (te) desde una perspectiva transdiagnóstica. Los protocolos transdiagnósticos enfatizan los procesos esenciales subyacentes a los distintos te, son adecuados para el tratamiento de distintos trastornos psicológicos y permiten un abordaje más adecuado de la comorbilidad entre este tipo de trastornos. Por otra parte, el uso de las tecnologías de la información y la comunicación (tic) puede ayudar a que los tratamientos psicológicos lleguen a un mayor número de personas, contribuyendo de esta manera a mejorar su eficiencia. La mayoría de tra-bajos sobre protocolos de tratamiento online se centran en el estudio de su eficacia, sin embargo es importante también estudiar la aceptabilidad de este tipo de interven-ciones. Nuestro grupo de investigación ha desarrollado un protocolo de tratamiento transdiagnóstico para los te, y lo ha adaptado para que pueda ser aplicado a través de Internet. En este trabajo se describe este protocolo y se presentan datos prelimi-nares sobre su aceptabilidad. metodología: 12 participantes (edad media = 28,58, d. t. = 5,35) de una muestra clínica con un diagnóstico de te que recibieron un pro-tocolo de tratamiento transdiagnóstico combinado (terapia presencial + protocolo de tratamiento online). Se obtuvieron datos acerca de las expectativas y opinión de los participantes evaluadas con la escala de expectativas sobre el tratamiento y la esca-la de opinión sobre el tratamiento. resultados: Las puntuaciones obtenidas fueron elevadas en ambas variables relacionadas con la aceptación (expectativas y opi-nión). discusión: El estudio de la aceptabilidad por parte de los pacientes hacia estos programas es importante, ya que puede influir en la decisión acerca de iniciar y/o seguir este tipo de tratamientos. Contar con esta información puede ayudarnos en el desarrollo y mejora de los tratamientos psicológicos autoaplicados a través de Internet. Introduction: In recent years there has been great interest in addressing the treatment of ed from a transdiagnostic perspective. Transdiagnostic treatment protocols empha-size the essential underlying processes that are common to ed and are conceived of for the treatment of several mental disorders. Also, comorbidity among these disorders can be more adequately addressed from this perspective. In addition, Information and Communication Technologies (icts) can facilitate access by people for whom traditional therapy is not available thereby contributing to an improvement in its cost-effectiveness. Most of the articles about online treatment protocols are efficacy studies, nevertheless it is also important to analyze the acceptability of these types of interventions. Our re-search group has developed a transdiagnostic treatment protocol for the treatment of ed that has been adapted to be applied online over the Internet. The aim of this article is to describe the treatment protocol and to present data about its acceptability. me-thodology: 12 participants (mean age = 28.58, d. t. = 5.35) from a clinical sample with a diagnosis of ed which received a blended transdiagnostic protocol treatment (face-to-face therapy plus online treatment protocol). Scores of expectancies and opinion of treatment were assessed with the Expectancies Treatment Scale and the Opinion Treatment Scale. results: High scores in both variables related to acceptability (expec-tations and opinion of treatment) were obtained. discussion: The study of the accep-tability towards this kind of programs is important, since it may influence the participant’s decision to initiate and/or follow this kind of treatments. Counting on this information can help us in the development and improvement of Internet-based treatment proto-cols.

Some structural observations on cultured Vahlkampfia sp. trophozoites are reported. Trophozoites are active and pleomorphic, producing large cell protrusions related to locomotion such as lamellipodia, filopodia and endocytic structures formed by hyaline cytoplasm, in which actin provides a framework that allows rapid changes in morphology. As observed by transmission electron microscopy, the cytoplasm is highly granular masking some cell organelles and the major cytoplasmic membrane systems. The structure of cell organelles such as the nucleus, endoplasmic reticulum, and digestive vacuoles is described. A common finding was the presence of 50 nm electron-dense round granules that are not limited by a membrane and that appear scattered in the cytoplasm, and whose function remains unknown. Apparently, the cell reserve material is glycogen, since complete trophozoites were positive to Schiff periodic-acid technique.

The effect of chloroform extracts from seeds of the papaya plant (Carica papaya) on the spermatic characteristics of dogs was evaluated at doses of 50, 100 and 150 mg/kg, Groups 1, 2, and 3, respectively. Structural and ultrastructural changes in sperm cells and testicular parenchyma were also evaluated, and possible side effects were noted. Significant reductions in sperm concentration and motility were observed starting from Days 60 and 75, respectively, in all treated groups (P0.05), but no azoospermia was noted. A mild osmotic diarrhea occurred in dogs from Group 3 (150 mg/kg), although blood variables were within the normal range of a clinically healthy dog. Arrested spermatogenesis was observed in the seminiferous tubules of all treated groups, and vacuolization and signs of Sertoli cell degeneration were detected in all treated groups, particularly in Group 3 (150 mg/kg). Selective damage to Sertoli cells induced by the extract occurred in all treated groups independently of the extract concentration. Alteration of the epididymal environment may reduce the motility of sperm cells, considering that their structure was normal. Sperm characteristics in treated animals were considered to be similar to those of sub-fertile dogs. However, these effects may be temporary, and dogs may recover normal sperm characteristics when the extract is withdrawn.

Thyroid hormones play a major role in the maturation process of the brain. Currently, congenital hypothyroidism is detected by mass screening. The impact of this early hormonal deficiency on the organization of the sleep pattern is not known. In this study, the polysomnographic features in children diagnosed with congenital hypothyroidism were analyzed. Children were detected by mass population screening and the hormonal replacement therapy starts immediately. Children's age ranged between 1.5 and 18 months of age. The duration of hormonal treatment before sleep recordings varied between 8 days and 17 months. Children were polysomnographically recorded in the morning, for at least 2h, obtaining more than one sleep cycle. Results showed a high prevalence of females (5/1) in the group studied. A high proportion of infants (43%) displayed central apnea in different degrees (mild, moderate and severe) as well as hypopnea (83%), mainly in subjects around 4 and 8 months of age. The proportion of infants displaying central apnea decreases as age increases. In addition, indeterminate (light) sleep increase and quiet (slow wave) sleep decrease significantly regardless of age and treatment. The percentage of REM sleep correlated positively with the age of the child at the beginning of the treatment, and negatively with their age at the time of the study. These data indicate that congenital hypothyroidism facilitates the presence of central sleep apnea. The decrease of these respiratory alterations correlates with the increase of the hormonal replacement therapy. It seems that sleep respiratory alterations in congenital hypothyroidism are linked to brain maturation processes in which thyroid hormones play a major role.

DAPI and Feulgen stains were used as specific DNA markers for studying the mitosis process in Naegleria fowleri. Both DAPI and Feulgen stains reacted with DNA in the nuclei of the amoebae. Representative figures of N. fowleri mitotic nuclei with a defined arrangement according to the phase of the cell cycle were observed. A notable characteristic is that the nucleolus is present throughout the stages of mitosis. During metaphase, several deeply stained DNA condensations following an elongated pattern were observed, corresponding almost certainly to tightly grouped chromosomes. Ultrastructural observations demonstrated that the nucleus divides by cryptomitosis, a process in which the nuclear membrane does not disappear during the mitosis. Centrioles were not found, and a spindle of microtubules was observed running the length of the nucleus from pole to pole however, they did not come to a focal point.

Free-living amoebae (FLA) are widely distributed worldwide. Some genera included in this group act as opportunistic pathogens causing fatal encephalitis and Acanthamoeba keratitis (AK), a sight-threatening infection of the cornea associated with the use of soft contact lenses that could even end in blindness if an early diagnosis and treatment are not achieved. Furthermore, the numbers of AK cases keep rising worldwide mainly due to an increase of contact lens wearers and lack of hygiene in the maintenance of lenses and their cases. In Mexico, no cases of AK have been described so far although the isolation of other pathogenic FLA such as Naegleria fowleri and Balamuthia mandrillaris from both clinical and environmental sources has been reported. The present study reports two cases of Acanthamoeba keratitis diagnosed in two patients admitted to the Hospital “Luis Sanchez Bulnes” for Blindness Prevention in Mexico City, Mexico. Corneal scrapes and contact lenses were checked for the presence of Acanthamoeba strains in both patients. Strains were axenized after initial isolation to classify at the genotype level. After sequencing the diagnostic fragment 3 (DF3) region located on the 18S ribosomal DNA (rDNA) gene of Acanthamoeba, genotype T3 and genotype T4 were identified in clinical case 1 and 2, respectively. To our knowledge, these are the first reported cases of AK in Mexico in the literature and the first description of Acanthamoeba genotypes T3 and T4 as causative agents of amoebic infection.

The fermentative and aromatic capabilities of Kloeckera africana/Hanseniaspora vineae K1, K. apiculata/H. uvarum K2, and Saccharomyces cerevisiae S1 and S2 were studied in pure and mixed culture fermentations using Agave tequila juice as the culture medium. In pure and mixed cultures, Kloeckera/Hanseniaspora strains showed limited growth and sugar consumption, as well as low ethanol yield and productivity, compared to S. cerevisiae, which yielded more biomass, ethanol and viable cell concentrations. In pure and mixed cultures, S. cerevisiae presented a similar behaviour reaching high biomass production, completely consuming the sugar, leading to high ethanol production. Furthermore, the presence of S. cerevisiae strains in the mixed cultures promoted the production of higher alcohols, acetaldehyde and ethyl esters, whereas Kloeckera/Hanseniaspora strains stimulated the production of ethyl acetate and 2-phenyl ethyl acetate compounds.

Entamoeba histolytica trophozoites are able to degrade human erythrocytes; the loss of erythrocyte cellular matrix and degradation of plasma membrane were observed, along with the decrease in the average size of digestive vacuoles. Ninety-six percent of hemoglobin ingested was hydrolyzed by trophozoites within 3 h, as evidenced by electrophoresis. Accordingly, X-ray spectroscopy revealed the presence of iron inside vacuoles after erythrophagocytosis, the concentration of which decreased to control levels in a similar period. Quantification of erythrocyte digestion at the early and late periods was determined by a spectrophotometric procedure, with t1/2 = 1.67 h and 35-min for HM-1:IMSS and HK-9:NIH trophozoites, respectively. In the latter, activity was due to the combined action of intracellular enzimatic activity and exocytosis. E-64c and leupeptin totally inhibited erythrocyte digestion within a 3-h period, thereafter hydrolysis took place at lower rate. Our results suggest that erythrocyte digestion in E. histolytica proceeds in different ways in these two amebic strains, and can be blocked by proteinase inhibitors.

Specific anti-Acanthamoeba IgA antibodies have been detected in the serum and tears of patients and healthy individuals. However, the role of human secretory IgA antibodies in inhibiting the adherence of Acanthamoeba had not been previously investigated. Therefore, the purpose of this study was to purify secretory IgA from human colostrum and analyze its effect on the adherence of Acanthamoeba trophozoites to contact lenses and Madin–Darby canine kidney (MDCK) cells. IgA antibodies to Acanthamoeba polyphaga in colostrum of healthy women as well as in saliva and serum of healthy subjects were analyzed by ELISA and Western blot analysis. In serum, saliva, and colostrum, we detected IgA antibodies that recognized several antigens of A. polyphaga. In addition, colostrum and IgA antibodies purified from it inhibited adherence of A. polyphaga trophozoites to contact lenses and MDCK cells. These results suggest that IgA antibodies may participate in the resistance to the amoebic infection, probably by inhibiting the adherence of the trophozoites to contact lenses and corneal epithelial cells.Key words: Acanthamoeba polyphaga, free-living amoebas, colostrum, IgA.

Background. The Trypanosomatidae family possesses one of the most unusual DNAs found in nature: the kinetoplast genome. It consists of a few dozen maxicircles that encode for some subunits of mitochondrial enzymes and rRNAs in a cryptic pattern and thousands of minicircles that encode for the guide RNAs (gRNAs), all catenated and constituting a dense network. The complexity of kinetoplast genome based on its intricate DNA structure is well known; however, only a small number of proteins associated with kinetoplast DNA (kDNA) have been described, and the majority are related with the replication process. Methods. We describe the protein profile obtained using formaldehyde as a cross-linking agent to obtain the kinetoplast DNA-protein complex, and Southwestern assay to identify the kDNA binding proteins present in the complex. Results. We identified seven proteins eluted from the kDNA complex fixed by formaldehyde. Polyclonal antiserum developed against the kDNA-protein complex recognized only four proteins in crude extracts of epimastigote stage, suggesting immunogenic differences among these proteins and/or their availability in the kinetoplast genome. Southwestern assay using minicircle fragments showed nine kDNA binding proteins in crude extracts of Trypanosoma cruzi epimastigote. Conclusions. We describe several proteins associated with the kDNA. Some could be involved in the essential process for parasite life and also could be a good target for drug or vaccine development. The results contribute to understanding the organization of the kinetoplast genome. © 2002 IMSS. Published by Elsevier Science Inc.

Con el objetivo de que los sistemas y subsistemas de educación media superior se integren al Sistema Nacional de Bachillerato (SNB), el Consejo para la Evaluación de la Educación del tipo Media Superior (COPEEMS) se encarga de evaluar y acreditar a las instituciones educativas de Educación Media Superior que tengan la intención de pertenecer al SNB. En 2014, COPEEMS recomendó la actualización de los cursos al Bachillerato General por Áreas Interdisciplinarias (BGAI) –modalidades mixta y virtual– de la Universidad de Guadalajara (UDG). El presente trabajo describe y analiza la experiencia de actualización y rediseño de los programas de estudio de dicho bachillerato.

Alzheimer's disease (AD) is the most common cause of dementia in the elderly. AD brains are characterized by the presence of neurofibrillary tangles (NFTs) and neuritic plaques. NFTs are constituted of paired helical filaments, which are structurally composed by assembled hyperphosphorylated and truncated tau polypeptides. To date, the integral constituents of NFTs remain unknown mainly due to the high insolubility of NFTs. The aim of this study was to identify by tandem mass spectrometry, the polypeptides contained in both isolated NFTs by laser capture microdissection and total homogenates, using tissue sections from paraformaldehyde-fixed AD brains. In the first case, we isolated 2,000 NFTs from tissue samples of hippocampus from each of the three Mexican AD brains used in our study. These were previously stained with anti-hyperphosphorylated tau AT-100 antibodies. After the removal of paraformaldehyde and delipidation with organic solvents, we tested three solubilization methods. We identified 102 polypeptides from total homogenates and 41 from isolated NFTs. We selected UCH-L1, transferrin, and GAPDH polypeptides to be studied by immunofluorescence and confocal microscopy. Only UCH-L1 and GAPDH colocalized with hyperphosphorylated tau in NFTs.

The morphological analysis of the cytopathic effect on MDCK cell monolayers and hamster cornea and qualitative and quantitative analyses of conditioned medium and proteases were evaluated and compared between two strains ofAcanthamoebagenotype T4. Further than highlighting the biological differences found between both strains, the most important observation in this study was the fact that proteases both in total extracts and in conditioned medium are apparently not determinant in tissue destruction. An interestingly finding was that no lysis of corneal tissue was observed as it was previously suggested. These results, together with previous studies, allow us to conclude that the invasion and disruption of corneal tissue is performed by the penetration of the amoebae through cell junctions, either by the action of proteases promoting cellular separation but not by their destruction and/or a mechanical effect exerted by amoebae. Therefore, contact-dependent mechanisms inAcanthamoebapathogenesis are more relevant than it has been previously considered. This is supported because the phagocytosis of recently detached cells as well as those attached to the corneal epithelium leads to the modification of the cellular architecture facilitating the migration and destruction of deeper layers of the corneal epithelium.

Four phages isolated from cattle and poultry feces were analyzed for their ability to lyse Salmonella serotypes and Escherichia coli O157:H7. The phage one-step growth curves, morphology, and genetic characteristics were determined. All phages showed a lytic effect on various Salmonella serotypes and E. coli O157:H7, which lysed at least 70% of the 234 strains tested. The phages had latent periods ranging from 10 to 15 min and generation times of 30 to 45 min, while burst size fluctuated between 154 and 426 PFU/cell. Phages morphology showed isometric and elongated heads and rigid contractile tails, consistent with morphology of the Myoviridae family. Phages’ DNA dendrograms showed a distinctive RFLP when digested by HindIII and EcoRV, and SDS–PAGE profile showed distinctive proteins expression as well. In vitro phage challenge showed a total reduction of E. coli O157:H7, Salmonella Typhimurium and Saintpaul counts at 2 h, whereas for Salmonella Montevideo a reduction and retardation growth, at a multiplicity of infection (MOI) of 100, was observed; however, under a MOI of 10 000, no viable cells were detected after 4 h. The wide host ranges of these phages suggested they could be used for simultaneous biocontrol of some Salmonella serotypes and E. coli O157:H7.

The study of the encystation process of Entamoeba histolytica has been hampered by the lack of experimental means of inducing mature cysts in vitro. Previously we have found that cytoplasmic vesicles similar to the encystation vesicles of Entamoeba invadens are present in E. histolytica trophozoites only in amebas recovered from experimental amebic liver abscesses. Here we report that a monoclonal antibody (B4F2) that recognizes the cyst wall of E. invadens also identifies a 48 kDa protein in vesicles of E. histolytica trophozoites recovered from hepatic lesions. This protein is less expressed in trophozoites continuously cultured in axenical conditions. As previously reported for E. invadens, the B4F2 specific antigen was identified as enolase in liver-recovered E. histolytica, by two-dimensional electrophoresis, Western blot and mass spectrometry. In addition, the E. histolytica enolase mRNA was detected by RT PCR. The antigen was localized by immunoelectron microscopy in cytoplasmic vesicles of liver-recovered amebas. The B4F2 antibody also recognized the wall of mature E. histolytica cysts obtained from human samples. These results suggest that the enolase-containing vesicles are produced by E. histolytica amebas, when placed in the unfavorable liver environment that could be interpreted as an attempt to initiate the encystation process.

The present study demonstrates that when Acanthamoeba castellanii trophozoites are co-cultivated with isolated human corneas, the amoeba can be invasive and cause damage to the intact corneal epithelium without the requirement of previous corneal abrasion. After adhesion, A. castellanii trophozoites migrate between cells forming bumps on the corneal cell layers and reaching Bowman s membrane in 3h, although no evidence of cell damage was observed until the phagocytic process was detected. Likewise, conditioned medium produced damage to the corneal cells that was proportional to the time of incubation, but this cytophatic effect involved only the most superficial layer of the human cornea and was not enough to explain amoebic invasion of Bowman s membrane. As a result of our observations, we suggest that the mechanical action of the trophozoites and phagocytosis of corneal cells during the process of corneal invasion are more important than previously suggested.

The presence of the cytoskeleton of Acanthamoeba castellanii was observed by means of cryo-electronmicroscopy and immunofluorescence techniques. This structure is formed largely by fibers and networks of actin located mainly in cytoplasmic locomotion structures as lamellipodia and as well as in various endocytic structures. In addition, the comparision between total actin content in whole extracts among different amoebae was made. The molecular weight of actin in A. castellanii was 44 kDa, and 45 kDa for Naegleria fowleri and Entamoeba histolytica.

Cysts represent a phase in the life cycle of biphasic parasitic protozoa that allow them to survive under adverse environmental conditions. Two events are required for the morphological differentiation from trophozoite to cyst and from cyst to trophozoite: the encystation and excystation processes. In this paper, we present a review of the ultrastructure of the encystation and excystation processes in Entamoeba invadens, Acanthamoeba castellanii, and Giardia lamblia. The comparative electron microscopical observations of these events here reported provide a morphological background to better understand recent advances in the biochemistry and molecular biology of the differentiation phenomena in these microorganisms.

Differences in the distribution of diverse cell surface coat markers were found between Naegleria fowleri and Naegleria lovaniensis. The presence of carbohydrate-containing components in the cell coat of the two species was detected by selective staining with ruthenium red and alcian blue. Using both markers, N. fowleri presented a thicker deposit than N. lovaniensis. The existence of exposed mannose or glucose residues was revealed by discriminatory agglutination with the plant lectin Concanavalin A. These sugar residues were also visualized at the cell surface of these parasites either by transmission electron microscopy or by fluorescein-tagged Concanavalin A. Using this lectin cap formation was induced only in N. fowleri. The anionic sites on the cell surface detected by means of cationized ferritin were more apparent in N. fowleri. Biotinylation assays confirmed that even though the two amoebae species have some analogous plasma membrane proteins, there is a clear difference in their composition.

In this study we report observations on the structural mechanisms of the cytopathic eVect of Acanthamoeba castellanii trophozoites on cultured MDCK cell monolayers. Co-incubations were carried out for a maximum of 24 h. The Wrst evidence of damage to the cell monolayer was detected by measuring the transepithelial resistance of cell monolayers that interacted with the amoebae. At 6 h, transepithelial resistance diminished to 51% and amoebae required 5–6 h to produce evidence of structural injury at the light microscopy level. Following 12 h of incubation, the cell monolayer was severely damaged. After making intimate contact with the surface of target cells, trophozoites detached cells from the substrate, lysed and by means of food-cups ingested the damaged cells. There was no morphological evidence of modiWcations in MDCK cell membranes, membrane fusion or junction formation between the amoeba and host plasma membrane. The lytic capacity of the amoebas appears to be the result of cytotoxic factors secreted by the amoebae since, when monolayers were incubated with conditioned medium, there was also a decrease in the transepithelial resistance. Besides, mechanical injury produced by the attachment and movement of the trophozoites may contribute to the disruption of the cell monolayer. As in other pathogenic amoebae, the cytopathic action of A. castellanii on the cell monolayers can subjectively be separated into four stages: adhesion, cytolysis, phagocytosis, and intracellular degradation.

Ultrastructural studies on Entamoeba histolytica have been carried out mostly with trophozoites cultured for many years. Under these conditions, the availability of nutrients and the absence of environmental stimuli may switch off some phenotypic characteristics of the parasite. As a result, virulence of E. histolytica diminishes with prolonged culture passages, and the ability to form cysts disappears in axenically maintained trophozoites. The present analysis by transmission electron microscopy of trophozoites recovered from experimental amebic liver lesions in hamsters revealed two types of cytoplasmic changes. On the one hand, the number of peripheral electron dense granules significantly increased in amebas obtained from liver lesions 15 min and 6h after inoculation. On the other hand, large cytoplasmic vesicles with a microfibrillar content appeared in trophozoites cultured from 72 or 96 h hepatic lesions. With fluorescence microscopy, a chitin-like material was identified in these vesicles by reactivity with calcofluor M2R. Ultrastructurally, these cytoplasmic components resemble the encystation vesicles of Entamoeba invadens and Giardia lamblia. The release of large amounts of electron dense granules, known to contain collagenase activity, probably contributes to degrade extracellular matrix components during tissue invasion. In addition, under the conditions mentioned above, amebas form encystation-like vesicles in an incomplete process of differentiation into cysts, which are the resistant form of the parasite.

In recent years, the use of fast-freeze fixation followed by freeze-substitution has been shown to be the procedure that best satisfies the ultrastructural preservation of cellular components due to the rapidity of the fixation procedure and the reduction of artifacts compared to chemical fixation. When these techniques were used to study the fine structure of axenically cultured trophozoites of Acantamoeba castellanii, an improved preservation of the whole cell was observed. The ground substance of the cytoplasm is densely packed with fibrogranular material which is frequently removed with the use of conventional techniques. Also, the ultrarapid physical stabilization allows the visualization of fusion and fission processes of cytoplasmic vacuoles and vesicles. The nuclear structure and cytoplasmic microfilaments as well as membranous structures were clearly identified. Low temperature techniques combine the advantage of fast-freeze fixation for the physical stabilization of organic molecules and their stabilization by the substitution medium at low temperature giving rise not only to a better preservation of the cell ultrastructure but providing a favorable basis for immunocytochemistry at the electron microscopy level.

Background The cell wall of Entamoeba invadens cysts is composed of chitin microfibrils as the main structural component. It has been demonstrated in yeast that the chitin cell wall assembly is altered by dyes such as Congo red (CR) and Calcofluor. Methods The purpose of this work was to study the cell wall assembly under the effect of CR dye on encysting E. invadens by means of light and electron microscopy, after the amebas were subjected to the effect of 100–2,000 μg CR/mL. Experiments were performed either in BI-S-33 or in mLG media. Results Trophozoite growth was not inhibited by 100–1,000 μg/mL CR after 8 days of incubation in BI-S-33 medium. However, low levels of growth were observed with 2,000 μg/mL of dye. No significant differences in morphologically viable (hyaline) cyst production occurred after 24–48 h, when 100 μg CR/mL was used, while the highest concentration of CR (2,000 μg/mL) resulted in a significant decrease of hyaline cyst yield; dead cysts prevailed in cultures, particularly at 72 h of CR treatment. Differentiation of amebas incubated in the presence of 500–2,000 μg/mL CR produced abnormal chitin deposits, rendering irregularly thick or double cell walls, as shown by transmission and scanning electron microscopy. Cyst cultures obtained under 100 μg/mL CR produced as many trophozoites as did the control when they were incubated in BI-S-33, but only low numbers of trophozoites were found in culture cysts obtained under higher CR doses. Conclusion Our results suggest that CR affects E. invadens encystment, alters the cell wall formation, and also affects the cyst viability.

This paper explores the interaction of two strains of Trichomonas vaginalis, of high and low virulence, with the cell types present in the microenvironment of the parasite during human infections. With the use of transmission and scanning electron microscopy the sequence of internalization by T. vaginalis of Doderlein's lactobacilli, and of vaginal epithelial cells, leukocytes, and erythrocytes was documented. Furthermore, the degradation of ingested material by colocalization of acid phosphatase activity in phagocytic vacuoles was demonstrated. Phagocytosis of all cell types analyzed was found in both strains studied, although the highly virulent strain internalized target cells more rapidly than the less virulent one. Ultrastructural evidence indicated that phagocytosis takes place through two distinct mechanisms, only one involving the formation of a phagocytic stoma, characteristic of professional phagocytes. T. vaginalis phagocytosis may be both an efficient means of obtaining nutrients for the parasite and a significant factor in the pathogenesis of trichomonal infections of the human genitourinary tract.

The surface charge of Giardia lamblia trophozoites from axenic cultures of strains recently isolated in Mexico from human cases of symptomatic and asymptomatic giardiasis was studied by means of cellular microelectrophoresis and ultrastructural cytochemistry. It is concluded that ionogenic surface groups confer a negative surface charge on trophozoites of G. lamblia and that no significant differences exist between the surface charge of trophozoites of symptomatic and asymptomatic origin.

THE study of the cellular properties of Entamoeba histolytica has been facilitated by the development of cultures under axenic conditions1. Various strains of E. histolytica have been isolated from patients with acute dysentery and from asymptomatic carriers. Because of the possible differences in the degree of virulence between these two types of strain, we studied their surface properties, using various methods which have been of value in identifying differences between normal and tumour cell surfaces2.