Avirulent UG10 Entamoeba histolytica mutant derived from HM-1:IMSS strain shows limited genome variability and aberrant 5-methyl cytosine genomic distribution.

Entamoeba histolytica, an intestinal parasite of global significance, poses substantial health risks with its associated high morbidity and mortality rates. Despite the current repertoire of molecular tools for the study of gene function in, the regulatory mechanisms governing its pathogenicity remain largely unexplored. This knowledge gap underscores the need to elucidate key genetic determinants orchestrating cellular functions critical to its virulence. Previously, our group generated an avirulent strain, termed UG10, with the same genetic background as the HM1:IMSS strain. UG10 strain, despite showing normal expression levels of well-known virulence factors, was unable to perform in-vitro and in-vivo activities related to amoebic virulence. In this study, we aimed to uncover the genome-wide modifications that rendered the avirulent phenotype of the UG10 strain through whole-genome sequencing. As a complementary approach, we conducted Methylated DNA Immunoprecipitation coupled with sequencing (MeDIP-seq) analysis on both the highly virulent HM1:IMSS strain and the low-virulence UG10 strain to uncover the genome-wide methylation profile. These dual methodologies revealed two aspects of the UG10 avirulent strain. One is the random integration of fragments from the ribosomal gene cluster and tRNA genes, ranging from 120 to 400 bp; and secondly, a clear, enriched methylation profile in the coding and non-coding strand relative to the start codon sequence in genes encoding small GTPases, which is associated with the previously described avirulent phenotype. This study provides the foundation to explore other genetic and epigenetic regulatory circuitries in E. histolytica and novel targets to understand the pathogenic mechanism of this parasite.

Theoretical study of ArcB and its dimerization, interaction with anaerobic metabolites, and activation of ArcA.

The complex metabolism of Escherichia coli has been extensively studied, including its response to oxygen availability. The ArcA/B two-component system (TCS) is the key regulator for the transition between these two environmental conditions and has been thoroughly characterized using genetic and biochemical approaches. Still, to date, limited structural data is available. The breakthrough provided by AlphaFold2 in 2021 has brought a reliable tool to the scientific community for assessing the structural features of complex proteins. In this report, we analyzed the structural aspects of the ArcA/B TCS using AlphaFold2 models. The models are consistent with the experimentally determined structures of ArcB kinase. The predicted structure of the dimeric form of ArcB is consistent with the extensive genetic and biochemical data available regarding mechanistic signal perception and regulation. The predicted interaction of the dimeric form of ArcB with its cognate response regulator (ArcA) is also consistent with both the forward and reverse phosphotransfer mechanisms. The ArcB model was used to detect putative binding cavities to anaerobic metabolites, encouraging testing of these predictions experimentally. Finally, the highly accurate models of other ArcB homologs suggest that different experimental approaches are needed to determine signal perception in kinases lacking the PAS domain. Overall, ArcB is a kinase with features that need further testing, especially in determining its crystal structure under different conditions.

Hiperglucemia en ayuno, medicamentos y plantas medicinales en un laboratorio clínico de Oaxaca, México / Fasting hyperglycemia, drugs and medicinal plants in a clinical laboratory in Oaxaca, Mexico

Resumen Objetivo: Determinar la prevalencia de hiperglucemia en ayuno y el uso de medicamentos y plantas medicinales en pacientes atendidos en el laboratorio de análisis clínicos HECA LAB de Ciudad Ixtepec, Oaxaca. Materiales y Métodos: Se realizó un estudio descriptivo, transversal durante el cuarto trimestre del año 2021. Se llevó a cabo un muestreo no probabilístico, por conveniencia, incluyendo un total de 205 pacientes. La recolección de los datos se llevó a cabo mediante la aplicación de un cuestionario estructurado que contenía información sociodemográfica. A los pacientes con hiperglucemia se les interrogó sobre el consumo de medicamentos y plantas medicinales. Los niveles de glucosa fueron determinados mediante el analizador semiautomático de química clínica Mindray, modelo BA-88A. Se consideró como diabético un valor de glucosa mayor o igual a 126 mg/dl de acuerdo a los criterios de la Organización Mundial de la Salud. Se calculó la prevalencia por fórmula, según Aschengrau & Seage. Resultados: De los 205 pacientes, el 29% presentó diabetes. El 8% se clasificó como prediabético con predominio del sexo femenino. El rango de edad más frecuente fue de 54-65 años con el 32%. El 82% refirió llevar tratamiento farmacológico y el 63% de los encuestados manifestó consumir plantas medicinales para el tratamiento de la diabetes. Conclusiónes: La prevalencia de diabetes en la muestra estudiada fue del 29%. La monoterapia oral con metformina, fue el tratamiento más frecuente, seguido por el uso de insulina vía subcutánea. Fueron referidas 11 plantas medicinales, siendo la moringa (Moringa oleifera) la más empleada. El 18% de los pacientes con diabetes no recibió tratamiento farmacológico. Por ello, es necesario establecer intervenciones oportunas de educación en salud, sobre cambios en el estilo de vida y adherencia al tratamiento farmacológico de los pacientes con diabetes mellitus; para lograr un manejo adecuado de la enfermedad y evitar complicaciones.

Abstract Objetive: To determinate the prevalence of fasting hyperglycemia and the use of drugs and medicinal plants in patients trated at the HECA LAB clinical analysis laboratory in Ciudad Ixtepec, Oaxaca. Materials and Methods: A descriptive, cross-sectional study was carried out during the fourth quarter of 2021. A non-probabilistic sampling was carried out, for convenience, incluiding a total of 205 patients. Data collection was carried out by applying a structured questionnaire containing sociodemographic information such as age and sex. Patients with hyperglycemia were questioned about the consumption of medicines and medicinal plants. Glucose levels were determined using the Mindray semi-automatic clinical chemistry analyzer, model BA-88A. A glucose value greater than or equal to 126 mg/dl according to the WHO criteria was considered diabetic. Prevalence was calculated by formula, according to Aschengrau & Seage. Results: Of the 205 patients, 29% had diabetes. 8% were classified as prediabetic with a predominance of females. The most frequent age range was 54-65 years with 32%. 82% reported taking pharmacological treatment and 63% of those surveyed reported consuming medicinal plantas for the treatment of diabetes. Conclusions: The prevalence of diabetes in the studied patients was 29%. Oral monotherapy with metformin was the most frequent treatment, followed by the use of subcutaneous insulin. Eleven medicinal plants were referred, being moringa (Moringa oleifera), the most used. 18% did not receive pharmacological treatment. For this reason, it is necessary to establish timely health education interventions on lifestyle changes and adherence to pharmacological treatment of patients with diabetes mellitus; to achieve adequate management of the disease and avoid complications.

Attenuation of In Vitro and In Vivo Virulence Is Associated with Repression of Gene Expression of AIG1 Gene in Entamoeba histolytica.

Entamoeba histolytica virulence results from complex host-parasite interactions implicating multiple amoebic components (e.g., Gal/GalNAc lectin, cysteine proteinases, and amoebapores) and host factors (microbiota and immune response). UG10 is a strain derived from E. histolytica virulent HM-1:IMSS strain that has lost its virulence in vitro and in vivo as determined by a decrease of hemolytic, cytopathic, and cytotoxic activities, increased susceptibility to human complement, and its inability to form liver abscesses in hamsters. We compared the transcriptome of nonvirulent UG10 and its parental HM-1:IMSS strain. No differences in gene expression of the classical virulence factors were observed. Genes downregulated in the UG10 trophozoites encode for proteins that belong to small GTPases, such as Rab and AIG1. Several protein-coding genes, including iron-sulfur flavoproteins and heat shock protein 70, were also upregulated in UG10. Overexpression of the EhAIG1 gene (EHI_180390) in nonvirulent UG10 trophozoites resulted in augmented virulence in vitro and in vivo. Cocultivation of HM-1:IMSS with E. coli O55 bacteria cells reduced virulence in vitro, and the EhAIG1 gene expression was downregulated. In contrast, virulence was increased in the monoxenic strain UG10, and the EhAIG1 gene expression was upregulated. Therefore, the EhAIG1 gene (EHI_180390) represents a novel virulence determinant in E. histolytica.

Escherichia coli transcription factors of unknown function: sequence features and possible evolutionary relationships.

Organisms need mechanisms to perceive the environment and respond accordingly to environmental changes or the presence of hazards. Transcription factors (TFs) are required for cells to respond to the environment by controlling the expression of genes needed. Escherichia coli has been the model bacterium for many decades, and still, there are features embedded in its genome that remain unstudied. To date, 58 TFs remain poorly characterized, although their binding sites have been experimentally determined. This study showed that these TFs have sequence variation at the third codon position G+C content but maintain the same Codon Adaptation Index (CAI) trend as annotated functional transcription factors. Most of these transcription factors are in areas of the genome where abundant repetitive and mobile elements are present. Sequence divergence points to groups with distinctive sequence signatures but maintaining the same type of DNA binding domain. Finally, the analysis of the promoter sequences of the 58 TFs showed A+T rich regions that agree with the features of horizontally transferred genes. The findings reported here pave the way for future research of these TFs that may uncover their role as spare factors in case of lose-of-function mutations in core TFs and trace back their evolutionary history.

Histochemical Evidence for Nitrogen-Transfer Endosymbiosis in Non-Photosynthetic Cells of Leaves and Inflorescence Bracts of Angiosperms.

We used light and confocal microscopy to visualize bacteria in leaf and bract cells of more than 30 species in 18 families of seed plants. Through histochemical analysis, we detected hormones (including ethylene and nitric oxide), superoxide, and nitrogenous chemicals (including nitric oxide and nitrate) around bacteria within plant cells. Bacteria were observed in epidermal cells, various filamentous and glandular trichomes, and other non-photosynthetic cells. Most notably, bacteria showing nitrate formation based on histochemical staining were present in glandular trichomes of some dicots (e.g., Humulus lupulus and Cannabis sativa). Glandular trichome chemistry is hypothesized to function to scavenge oxygen around bacteria and reduce oxidative damage to intracellular bacterial cells. Experiments to assess the differential absorption of isotopic nitrogen into plants suggest the assimilation of nitrogen into actively growing tissues of plants, where bacteria are most active and carbohydrates are more available. The leaf and bract cell endosymbiosis types outlined in this paper have not been previously reported and may be important in facilitating plant growth, development, oxidative stress resistance, and nutrient absorption into plants. It is unknown whether leaf and bract cell endosymbioses are significant in increasing the nitrogen content of plants. From the experiments that we conducted, it is impossible to know whether plant trichomes evolved specifically as organs for nitrogen fixation or if, instead, trichomes are structures in which bacteria easily colonize and where some casual nitrogen transfer may occur between bacteria and plant cells. It is likely that the endosymbioses seen in leaves and bracts are less efficient than those of root nodules of legumes in similar plants. However, the presence of endosymbioses that yield nitrate in plants could confer a reduced need for soil nitrogen and constitute increased nitrogen-use efficiency, even if the actual amount of nitrogen transferred to plant cells is small. More research is needed to evaluate the importance of nitrogen transfer within leaf and bract cells of plants.

Chromogenic Escherichia coli reporter strain for screening DNA damaging agents.

The presence of pollutants in soil and water has given rise to diverse analytical and biological approaches to detect and measure contaminants in the environment. Using bacterial cells as reporter strains represents an advantage for detecting pollutants present in soil or water samples. Here, an Escherichia coli reporter strain expressing a chromoprotein capable of interacting with soil or water samples and responding to DNA damaging compounds is validated. The reporter strain generates a qualitative signal and is based on the expression of the coral chromoprotein AmilCP under the control of the recA promoter. This strain can be used simply by applying soil or water samples directly and rendering activation upon DNA damage. This reporter strain responds to agents that damage DNA (with an apparent detection limit of 1 µg of mitomycin C) without observable response to membrane integrity damage, protein folding or oxidative stress generating agents, in the latter case, DNA damage was observed. The developed reporter strain reported here is effective for the detection of DNA damaging agents present in soils samples. In a proof-of-concept analysis using soil containing chromium, showing activation at 15.56 mg/L of Cr(VI) present in soil and leached samples and is consistent with Cr(III) toxicity at high concentrations (130 µg). Our findings suggest that chromogenic reporter strains can be applied for simple screening, thus reducing the number of samples requiring analytical techniques.

CLEAR - Medical use of contact lenses.

The medical use of contact lenses is a solution for many complex ocular conditions, including high refractive error, irregular astigmatism, primary and secondary corneal ectasia, disfiguring disease, and ocular surface disease. The development of highly oxygen permeable soft and rigid materials has extended the suitability of contact lenses for such applications. There is consistent evidence that bandage soft contact lenses, particularly silicone hydrogel lenses, improve epithelial healing and reduce pain in persistent epithelial defects, after trauma or surgery, and in corneal dystrophies. Drug delivery applications of contact lens hold promise for improving topical therapy. Modern scleral lens practice has achieved great success for both visual rehabilitation and therapeutic applications, including those requiring retention of a tear reservoir or protection from an adverse environment. This report offers a practical and relevant summary of the current evidence for the medical use of contact lenses for all eye care professionals including optometrists, ophthalmologists, opticians, and orthoptists. Topics covered include indications for use in both acute and chronic conditions, lens selection, patient selection, wear and care regimens, and recommended aftercare schedules. Prevention, presentation, and management of complications of medical use are reviewed.

Possible role played by the SINE2 element in gene regulation, as demonstrated by differential processing and polyadenylation in avirulent strains of E. histolytica.

Entamoeba histolytica represents a useful model in parasitic organisms due to its complex genomic organization and survival mechanisms. To counteract pathogenic organisms, it is necessary to characterize their molecular biology to design new strategies to combat them. In this report, we investigated a less-known genetic element, short interspersed nuclear element 2 (SINE2), that is present in this ameba and is highly transcribed and polyadenylated. In this study, we show that in two different nonvirulent strains of E. histolytica, SINE2 is differentially processed into two transcript fragments, that is, a full-length 560-nt fragment and a shorter 393-nt fragment bearing an approximately 18-nt polyadenylation tail. Sequence analysis of the SINE2 transcript showed that a Musashi-like protein may bind to it. Also, two putative Musashi-like sequences were identified on the transcript. Semiquantitative expression analysis of the two Musashi-like proteins identified in the E. histolytica genome (XP_648918 and XP_649094) showed that XP_64094 is overexpressed in the nonvirulent strains tested. The information available in the literature and the results presented in this report indicate that SINE2 may affect other genes, as observed with the epigenetic silencing of the G3 strain, by an antisense mechanism or via RNA-protein interactions that may ultimately be involved in the phenotype of nonvirulent strains of E. histolytica.

Etiologies and Features of Acute Viral Hepatitis in Spain.

BACKGROUND AND AIM: Etiologies of acute viral hepatitis in high-income countries change with migration of populations, lifestyle changes, and emergence of new pathogens. We analyzed etiologies, characteristics, and outcomes of patients with acute viral hepatitis at a tertiary hospital in Spain. METHODS: We analyzed data from all patients with acute hepatitis (n = 100; 71% male; median age, 42 years; 72% Spanish nationals), older than 16 years, diagnosed in the emergency department of an academic hospital in Barcelona, Spain, from January 2014 through December 2018. Blood samples were collected and patients with serum levels of alanine aminotransferase more than 10-fold the upper limit of normal and markers viral infection were considered to have acute viral hepatitis. We collected clinical information from patients, and samples were analyzed for IgM antibody to hepatitis B (HB) core antigen, HB surface antigen, antibody against hepatitis C virus (HCV), HCV RNA, IgM against hepatitis E virus (HEV), HEV RNA, and IgM against hepatitis A virus (HAV). Patients were followed until resolution of infections or evidence of chronic infection. RESULTS: The most common etiologies of acute hepatitis were HBV infection (28%), HEV infection (18%), HCV infection (17%), and HAV infection (14%). The main risk factors of the cohort were sexual risk contact and intravenous drug use; 79% of cases of HAV had sexual risk behavior. Twenty-nine percent of patients with acute HAV infection and 29% of patients with HBV infection were immigrants to Spain. Fifty-four patients were hospitalized; jaundice and HCV infection were associated with hospital admission. Three patients died (2 from acute liver failure related to acute HBV infection or HBV and HDV co-infection). Chronic infections developed in 5/28 patients (18%) with acute HBV infection and 7/17 patients (41%) with acute HCV infection. CONCLUSIONS: Despite universal vaccination against HBV in Spain, HBV remains the most frequent cause of acute viral hepatitis in our emergency department. Almost one-third of cases of acute HBV and HAV infections were immigrants, possibly from countries with suboptimal vaccination programs. A high proportion of patients with acute hepatitis have HEV infection (18%); acute HAV infection was associated with sexual risk behavior.

Cadet Agustín Melgar and military peripheral vascular trauma approach during Chapultepec's Battle.

Trauma to peripheral vascular organ is a very frequent condition during military conflicts. Fortunately, comprehensive understanding of local and systemic pathophysiology, in addition to the development of innovative surgical techniques and technological advances, have improved the outcome regarding to survival and anatomic and functional conservation of the limbs. In this manuscript, we perform an historical and state of the art review related to the approach of the peripheral vascular trauma, on the basis of an heroical episode from the Mexico City's Campaign, during North American invasion 1846-1848: the defense of Chapultepec's Castle by cadet Agustin Melgar.

El trauma del órgano vascular periférico es una condición muy frecuente en los conflictos militares. Por fortuna, la comprensión integral de la fisiopatología local y sistémica, en adición al desarrollo de técnicas quirúrgicas innovadoras y avances tecnológicos, ha mejorado el pronóstico relativo a la sobrevida y la conservación anatómica y funcional de las extremidades. En este trabajo se hace una revisión histórica y del estado actual del abordaje del trauma vascular periférico, sobre la base de un episodio heroico de la Campaña por la Ciudad de México durante la invasión norteamericana de 1846-1848: la defensa del Castillo de Chapultepec por el cadete Agustín Melgar.

aeBlue Chromoprotein Color is Temperature Dependent.

BACKGROUND: Marine sessile organisms display a color palette that is the result of the expression of fluorescent and non-fluorescent proteins. Fluorescent proteins have uncovered transcriptional regulation, subcellular localization of proteins, and the fate of cells during development. Chromoproteins have received less attention until recent years as bioreporters. Here, we studied the properties of aeBlue, a a 25.91 kDa protein from the anemone Actinia equina. OBJECTIVE: To assess the properties of aeBlue chromoprotein under different physicochemical conditions. METHODS: In this article, during the purification of aeBlue we uncovered that it suffered a color shift when frozen. We studied the color shift by different temperature incubation and physicochemical conditions and light spectroscopy. To assess the possible structural changes in the protein, circular dichroism analysis, size exclusion chromatography and native PAGE was performed. RESULTS: We uncover that aeBlue chromoprotein, when expressed from a synthetic construct in Escherichia coli, showed a temperature dependent color shift. Protein purified at 4 °C by metal affinity chromatography exhibited a pinkish color and shifts back at higher temperatures to its intense blue color. Circular dichroism analysis revealed that the structure in the pink form of the protein has reduced secondary structure at 4 °C, but at 35 °C and higher, the structure shifts to a native conformation and Far UV- vis CD spectra revealed the shift in an aromatic residue of the chromophore. Also, the chromophore retains its properties in a wide range of conditions (pH, denaturants, reducing and oxidants agents). Quaternary structure is also maintained as a tetrameric conformation as shown by native gel and size exclusion chromatography. CONCLUSION: Our results suggest that the chromophore position in aeBlue is shifted from its native position rendering the pink color and the process to return it to its native blue conformation is temperature dependent.

Plasma membrane damage repair is mediated by an acid sphingomyelinase in Entamoeba histolytica.

Entamoeba histolytica is a pathogen that during its infective process confronts the host defenses, which damages the amoebic plasma membrane (PM), resulting in the loss of viability. However, it is unknown whether amoebic trophozoites are able to repair their PM when it is damaged. Acid sphingomyelinases (aSMases) have been reported in mammalian cells to promote endocytosis and removal of PM lesions. In this work, six predicted amoebic genes encoding for aSMases were found to be transcribed in the HM1:IMSS strain, finding that the EhaSM6 gene is the most transcribed in basal growth conditions and rendered a functional protein. The secreted aSMase activity detected was stimulated by Mg+2 and inhibited by Co+2. Trophozoites that overexpress the EhaSM6 gene (HM1-SM6HA) exhibit an increase of 2-fold in the secreted aSMase activity. This transfectant trophozoites exposed to pore-forming molecules (SLO, Magainin, ß-Defensin 2 and human complement) exhibited an increase from 6 to 25-fold in the secreted aSMase activity which correlated with higher amoebic viability in a Ca+2 dependent process. However, other agents that affect the PM such as hydrogen peroxide also induced an increase of secreted aSMase, but to a lesser extent. The aSMase6 enzyme is N- and C-terminal processed. Confocal and transmission electron microscopy showed that trophozoites treated with SLO presented a migration of lysosomes containing the aSMase towards the PM, inducing the formation of membrane patches and endosomes in the control strain. These cellular structures were increased in the overexpressing strain, indicating the involvement of the aSMase6 in the PM injury repair. The pore-forming molecules induced an increase in the expression of EhaSM1, 2, 5 and 6 genes, meanwhile, hydrogen peroxide induced an increase in all of them. In all the conditions evaluated, the EhaSM6 gene exhibited the highest levels of induction. Overall, these novel findings show that the aSMase6 enzyme from E. histolytica promotes the repair of the PM damaged with pore-forming molecules to prevent losing cell integrity. This novel system could act when encountered with the lytic defense systems of the host.

Performance and predictability of a new large diameter contact lens design in keratoconic corneae.

OBJECTIVES: To evaluate the clinical performance and predictability using corneal topography in the fitting of a new large-diameter rigid gas permeable (RGP) contact lens design in eyes with keratoconus (KCN). METHODS: This study presents a review of eyes fitted with Alexa ES lenses for nonsurgical optimisation of visual correction. Anterior steep simulated keratometry (sim-K steep), corneal diameter (HVID), the Curvature at the apex in diopters (Cc), and distance from the corneal apex to the centre of the cornea (Lc) in millimetres derived from the Cone Location and Magnitude Index (CLMI) were recorded. Visual acuity, mean wearing time, final sagittal depth and adverse events were also recorded. Correlations between topographic indices and base curve were evaluated using the Pearson correlation coefficient. RESULTS: Forty-six eyes from 26 patients (19 males/ 7 females) were included. Mean visual acuity improved from 0.49 ± 0.32 with glasses, to -0.02 ± 0.10 with the contact lens. The mean daily wear time was 12.19 ± 1.96 hours. No complications were detected in 95.65% of the eyes [95% CI (83.9%-99.2%)] but two episodes of non-infectious keratitis. The average sagittal depth of the lenses fitted was 0.425 ± 0.15 mm, and it was positively correlated with the Cc value (r² = 0.66, p< 0.0001, n=46) derived from the CLMI index of the pre-fitting topography. CONCLUSIONS: Corneoscleral RGP contact lenses are a safe, and effective alternative for managing KCN patients to corneal lenses. Also, clinical data derived from the corneal topography could be used to help to decide the first diagnostic lens to be assessed, easing the overall fitting process.

Development of an oxidative stress sensor in live bacteria using the optimized HyPer2 protein.

Oxidative stress is a key regulator in many cellular processes but also an important burden for living organisms. The source of oxidative damage usually is difficult to measure and assess with analytical tools or chemical indicators. One major limitation is to discriminate the presence of secondary oxidant molecules derived from the cellular metabolism after exposure to the oxidant or the scavenging capacity of reactive oxygen species by cells. Using a whole-cell reporter system based on an optimized HyPer2 protein for Escherichia coli expression, we demonstrate that, as previously shown for eukaryotic organisms, the effect at the transcriptional level of hydrogen peroxide can be monitored in vivo using flow cytometry of bacterial cells without the need of a direct analytical measurement. In this approach, we generated two different HyPer2 expression systems, one that is induced by IPTG and a second one that is induced by oxidative stress responsive promoters to control the expression of the HyPer2 protein and the exposure of higher H2O2 concentrations that has been shown to activate oxidative response genes. Both systems showed that the pathway that leads to the generation of H2O2 in vivo can be traced from H2O2 exposure. Our results indicate that hydrogen peroxide pulses can be readily detected in E. coli cells by a defined fluorescence signature that is H2O2 concentration-dependent. Our findings indicate that although less sensitive than purified protein or expressed in eukaryotic cells, HyPer2 is a good bacterial sensor for H2O2. As proof of concept, this system was used to trace the oxidative capacity of Toluidine Blue O showing that oxidative stress and redox imbalance is generated inside the cell. This system is expanding the repertoire of whole cell probes available for tracing cellular stress in bacteria.

Repeatability of Anterior Curvature Metrics in Healthy and Keratoconic Eyes with a Portable Handheld Topographer.

SIGNIFICANCE: This is a prospective observational investigation that studies the repeatability of several outcomes obtained with the Keratron Scout in healthy and keratoconic corneas. In addition, we have been able to determine the "noise" values of the instrument, which can be very useful when evaluating possible progression in keratoconus (KCN). PURPOSE: The purpose of this study was to evaluate the repeatability of several anterior curvature metrics provided by a portable and handheld Placido disk-based topographer in healthy and keratoconic eyes. METHODS: One eye from 50 keratoconic patients and 50 eyes from healthy patients were included in this prospective observational study. Two consecutive sessions (with three scans per session) were performed, and the following parameters were analyzed with a Keratron Scout topographer: flat keratometry, steep keratometry, flat corneal axis, corneal astigmatism, and flat and steep asphericities. In addition, in the KCN cohort, indices derived from the Cone Location and Magnitude Index were also recorded. Intrasession (first session, all three measurements) and intersession (one measurement from each session, selected randomly) statistics were calculated. The following variables were calculated: within-subject standard deviation, coefficient of repeatability (R), coefficient of variation, and intraclass correlation coefficient (ICC). RESULTS: In healthy eyes, the intrasession and intersession ICCs were high (0.947 to 0.999), and for both flat keratometry and steep keratometry, the within-subject standard deviation was 0.08 and R was 0.24 diopters. Repeatability was slightly lower in the KCN group, but the ICC was greater than 0.95, whereas the R for flat keratometry was 0.41 diopters. CONCLUSIONS: The Keratron Scout provides repeatable measures for the studied metrics in healthy and keratoconic eyes.

Contribution of neutral sphingomyelinases to in vitro virulence of Entamoeba histolytica.

Amoebiasis is a worldwide health problem caused by the pathogen Entamoeba histolytica. Several virulence factors have been implicated in host invasion, immune evasion, and tissue damage. There are still new factors that remain to be elucidated and characterized. In this work, we obtained amoebic transfectants overexpressing three of the neutral sphingomyelinase enzymes encoded in the E. histolytica genome. The EhnSM3 overexpression induced an increase in hemolytic and cytotoxic activities, besides an increase in gene expression of amoebapore A, B, and C. Meanwhile the EhnSM1 and EhnSM2 overexpression caused an increase in cytopathic activity. In all the neutral sphingomyelinases overexpressing strains, the gene expression levels for cysteine proteinase 5, adhesin 112 and, heavy and light Gal/GalNAc lectin subunits were not affected. We propose that the increase of cytotoxic and lytic effect of EhnSM3 overexpressed strain can be related to the sum of the effect of EhnSM3 plus amoebapores, in a process cell contact-dependent or as mediator by inducing the gene expression of amoebapores enabling a link between EhnSM3 with the virulence phenotype in E. histolytica. Our results suggest a differential role for neutral sphingomyelinases in E. histolytica virulence.

[Toracotomía en el departamento de urgencias en una paciente con actividad eléctrica sin pulso no traumática: potencial ampliación del horizonte terapéutico de una intervención de alta especialidad]. / Emergency department thoracotomy in a patient with non-traumatic pulseless electrical activity: potential widening of the therapeutic horizon of a high-specialty intervention.

Instituida bajo recomendaciones objetivas, la toracotomía en el departamento de urgencias (TDU) se ha descrito como una maniobra quirúrgica salvatoria de la vida en pacientes traumatizados in extremis. Sin embargo, hay pocos reportes acerca de la experiencia con su empleo en la actividad eléctrica sin pulso no traumática. Describimos el caso de una paciente obstétrica exanguinada por sangrado masivo transoperatorio, en la que se realizó una TDU con un resultado óptimo para la vida y la función neurológica. Adicionalmente, evaluamos la literatura correspondiente al tema, que en lo mejor de nuestro conocimiento es crítica para expandir los protocolos de abordaje del ritmo de paro cardiaco no traumático en los hospitales de alto volumen.Instituted under objective recommendations, Emergency Department Thoracotomy (EDT) has been described as a life-saving surgical maneuver in trauma patients arriving "in extremis." Nevertheless, there are few reports related to the experience regarding the use of the procedure in non-traumatic cardiopulmonary arrest. We describe the case of an obstetric patient exsanguinated by operative massive bleeding, where EDT was instituted reaching an optimal result for the survival and neurologic function. Additionally, we evaluate the literature related to this issue, which to the best of our knowledge, is critical to expand protocols of approach in non-traumatic cardiac arrest rhythm in high-volume hospitals.

Red fluorescent protein (DsRFP) optimization for Entamoeba histolytica expression.

Entamoeba histolytica genetic organization and genome structure is complex and under intense research. The genome is fully sequenced, and several tools have been developed for the molecular study of this organism. Nevertheless, good protein tracking tags that are easy to measure and image, like the fluorescent proteins are lacking. In this report, we codon-optimized the red fluorescent protein from the coral Discosoma striata (DsRFP) for its use in E. histolytica and demonstrated functionality in vivo. We envision that this protein can be widely used for the development of transcriptional reporter systems and protein-tagging applications.