Evaluation of virulence factors in clinical isolates of pathogenic E. coli in avian samples in Caloto, Colombia

Avian pathogenic E. coli (APEC), produces an extraintestinal infection in chickens, turkeys, and other types of birds, called colibacillosis, which is considered one of the main causes of economic losses due to morbidity, mortality, and discard of poultry carcasses. The objective of the present study was to characterize the genetic profile of the virulence factors of different isolates of avian E. coli in Caloto, Cauca, Colombia. Materials and methods: E. coli was isolated and identified by biochemical tests, from 47 clinical isolates. Subsequently, the DNA was extracted using Chelex. Three multiplex PCRs were designed to amplify 13 virulence factors (iroN, hlyF, iss, iutA, frz, vat, sitA, KpsM, sitD, fimH, pstB, sopB, and uvrY), using primers previously reported for each. At the end, the amplification products were verified on agarose gels. Each isolate was classified according to the number of virulence factors: group A (between 10 and 13), group B (between 5 and 9), and group C (4 or less). Discussion and Conclusions: we were able to identify the presence of a group of virulence factors in clinical isolates of APEC, which allows us to demonstrate that both the frequency and the profile of virulence factors in the isolated strains showed a different profile than the reported by other authors. The virulence genes pstB and fimH were detected in all our samples, and the iss gene was the one with the lowest frequency. Finally, according to the number of virulence factors, the group A was the most frequent.

La E. coli patógena aviar (APEC), produce una infección extraintestinal en pollos, pavos y otros tipos de aves, denominada colibacilosis, la cual es considerada una de las principales causas de pérdidas económicas por morbilidad, mortalidad y descarte de canales de aves. El objetivo del presente estudio fue caracterizar el perfil genético de los factores de virulencia de diferentes aislamientos de E. coli aviar en Caloto, Cauca, Colombia. Materiales y métodos: E. coli se aisló e identificó mediante pruebas bioquímicas, a partir de 47 aislamientos clínicos. Posteriormente, el ADN se extrajo utilizando Chelex. Se diseñaron tres PCR multiplex para amplificar 13 factores de virulencia (iroN, hlyF, iss, iutA, frz, vat, sitA, KpsM, sitD, fimH, pstB, sopB y uvrY), utilizando primers informados previamente para cada uno. Al final, los productos de amplificación fueron verificados en geles de agarosa. Cada aislamiento se clasificó según el número de factores de virulencia: grupo A (entre 10 y 13), grupo B (entre 5 y 9) y grupo C (4 o menos). Discusión y Conclusiones: pudimos identificar la presencia de un grupo de factores de virulencia en los aislados clínicos de APEC, lo que nos permite demostrar que tanto la frecuencia como el perfil de los factores de virulencia en las cepas aisladas presentaron un perfil diferente al reportado por otros autores. Los genes de virulencia pstB y fimH se detectaron en todas nuestras muestras, siendo el gen iss el de menor frecuencia. Finalmente, según el número de factores de virulencia, el grupo A fue el más frecuente.

Crizotinib and erlotinib inhibits growth of c-Met+/EGFRvIII+ primary human glioblastoma xenografts.

OBJECTIVES: Receptor tyrosine kinases (RTK), such as c-Met and epidermal growth factor receptor (EGFR), are implicated in the malignant progression of glioblastoma. Studies show that RTK systems can co-modulate distinct and overlapping oncogenic downstream signaling pathways. EGFRvIII, a constitutively activated EGFR deletion mutant variant, leads to increased tumor growth and diminishes the tumor growth response to HGF: c-Met pathway inhibitor therapy. Conversely, activation of the c-Met pathway diminishes the tumor growth response to EGFR pathway inhibitors. Previously we reported that EGFRvIII and c-Met pathway inhibitors synergize to inhibit tumor growth in isogenic GBM cell lines engineered to express EGFRvIII. More recently, studies suggest that despite targeting RTK signaling in glioblastoma multiforme, a subpopulation of stem-like tumor-propagating cells can persist to replenish the tumor cell population leading to tumor recurrence. PATIENTS AND METHODS: Mayo 39 and Mayo 59 xenograft lines were cultured and xenografts were maintained. Subcutaneous xenograft lines were serially passaged in nude mice to generate subcutaneous xenografts. Xenografts were implanted in 6-8 week old nude mice. Once tumors reached a substantial size (150 mm3), mice were randomly divided into 4 groups: 1) control vehicle, 2) Crizotinib (crizo), 3) Erlotinib (erlot), or 4) Crizotinib + Erlotinib, (n = 5 per group). RESULTS: Crizotinib (c-Met pathway inhibitor) and Erlotinib (EGFR pathway inhibitor) in combination significantly inhibited tumor growth, phospho-EGFRvIII, phospho-Met, phospho-AKT, phospho-MAPK, and neurosphere growth in Mayo 39 and Mayo 59 primary GBM subcutaneous xenografts. The expression of the stem cell markers Nestin, Musashi, Olig 2 and Sox2 were also significantly down-regulated by c-Met inhibition, but no additive down-regulation was seen by co-treatment with Erlotinib. CONCLUSIONS: These results are consistent with and corroborate our previous findings demonstrating that targeting these two parallel pathways with c-Met and EGFR inhibitor therapy provides substantial anti-tumor activity in glioblastoma models.

Ultrasonographic ovarian dynamic, plasma progesterone, and non-esterified fatty acids in lame postpartum dairy cows.

The objective of this study was to compare ovulation rate, number of large ovarian follicles, and concentrations of plasma progesterone (P4) and non-esterified fatty acids (NEFA) between lame (n = 10) and non-lame (n = 10) lactating Holstein cows. The study was conducted in an organic dairy farm, and cows were evaluated by undertaking ultrasonography and blood sampling every 3 days from 30 days postpartum for a period of 34 days. Cows which became lame during the first 30 days postpartum experienced a lower ovulation rate determined by the presence of a corpus luteum (50% presence for lame cows and 100% for non-lame cows, p ≤ 0.05). The number of large ovarian follicles in the ovaries was 5 for lame cows and 7 for non-lame cows (p = 0.09). Compared to non-lame cows, lame cows had significantly lower (p ≤ 0.05) concentrations of plasma P4. Furthermore, NEFA concentrations were lower (p ≤ 0.05) in lame cows than in non-lame cows. It is concluded that lameness in postpartum dairy cows is associated with ovulation failure and lower concentrations of P4 and NEFA.

Adolph Lippe en la Historia de la Homeopatía / Adolph Lippe in the History of Homeopathy

Los fundamentos de la homeopatía fueron descritos y divulgados por Samuel Hahnemann y junto al maestro encontramos un grupo de seguidores fieles a sus enseñanzas, pioneros en difundir su legado; entre estos pioneros, Adolph Lippe reúne la mayor experiencia clínica exitosa documentada, demostrando así su destreza clínica gracias a la aplicación y a la defensa de los principios y de los fundamentos de la homeopatía, con total fidelidad a la ley de la semejanza, basándose en la Materia Médica Pura, en el Órganon y en el Tratado de Enfermedades Crónicas de Hahnemann. No obstante, en nuestro contexto poco se hace referencia a este eminente homeópata, desaprovechando la gran cantidad de información escrita que prueba la eficacia del método homeopático por él practicado. El objetivo de esta investigación fue definir el legado que dejó Adolph Lippe a la homeopatía. Se aplicó un método cualitativo, documental, descriptivo, no sistematizado y hermenéutico, donde se recolectó información de libros clásicos, sitios Web, tesis, artículos y otro tipo de documentos relacionados con las temáticas del estudio. Los resultados señalan que el legado de Lippe está constituido por: a) el ejemplo que dio del estudio constante y profundo de los tres libros (El Órganon, Las Enfermedades Crónicas y la Materia Médica Pura) que fundamentan el saber homeopático; b) su éxito clínico lo basó en lo descubierto por Hahnemann y nada más; c) sus escritos son un referente importante que nos recuerda hoy que el método hahnemanniano tiene toda su validez; d) la defensa de la homeopatía se hace con clínica exitosa. Un autor que ha aportado con esfuerzo y dedicación en la recuperación y la divulgación de la obra de Lippe, es el doctor André Saine.

The carboxyl-terminal tail of Noxa protein regulates the stability of Noxa and Mcl-1.

The BH3-only protein Noxa is a critical mediator of apoptosis and functions primarily by sequestering/inactivating the antiapoptotic Bcl-2 family protein Mcl-1. Although Noxa is a highly labile protein, recent studies suggested that it is degraded by the proteasome in a ubiquitylation-independent manner. In the present study, we investigated the mechanism of Noxa degradation and its ability to regulate the stability of Mcl-1. We found that the ubiquitylation-independent degradation of Noxa does not require a physical association with Mcl-1. A short stretch of amino acid residues in the C-terminal tail was found to mediate the proteasome-dependent degradation of Noxa. Ectopic placement of this degron was able to render other proteins unstable. Surprisingly, mutation of this sequence not only attenuated the rapid degradation of Noxa, but also stabilized endogenous Mcl-1 through the BH3-mediated direct interaction. Together, these results suggest that the C-terminal tail of Noxa regulates the stability of both Noxa and Mcl-1.

Experiencia local con el uso de punción única peribulbar caruncular para anestesia oftálmica / Local experience with caruncular single injection peribulbar anesthesia

Introducción: En la actualidad la anestesia peribulbar se considera la técnica de anestesia regional de elección para diversos procedimientos quirúrgicos oftalmológicos debido a su efectividad y a su baja incidencia de complicaciones. Técnicas recientes reducen las 2 punciones a una sola y se encuentran en proceso de evaluación. Objetivos:Medir la eficacia y la seguridad de la técnica de punción única peribulbar caruncular para distintos procedimientos quirúrgicos llevados a cabo en una clínica oftalmológica en la ciudad de Popayán (Colombia). Métodos: Se incluyeron pacientes sometidos a diversos procedimientos oftalmológicos. La técnica anestésica empleada consistía en una punción única peribulbar caruncular. El anestésico utilizado fue bupivacaína 0,5% 2,5 ml, lidocaína 2% 2,5 ml con hialuronidasa 7 UI/ml. Los pacientes fueron evaluados en los minutos 10,15 y 20. Se midió la funcionalidad motora de los 4 músculos extraoculares y el control motor del párpado superior e inferior. Resultados:Se incluyeron 137 pacientes. El 54% eran de género femenino y el 77% ASA II. A los 10 min de evaluación el 92% de los pacientes alcanzan anestésico apropiado para cirugía. Treinta y seis pacientes (26,3%) requirieron el uso de refuerzo mediante una nueva punción peribulbar. Veintidós pacientes (16%) refirieron presentar un dolor leve durante el procedimiento anestésico. En 4 casos (3%) se presentó quemosis. Conclusiones: Con el uso de la técnica de punción única caruncular la gran mayoría de los pacientes alcanzan un bloqueo ocular apropiado para diversos procedimientos quirúrgicos oftalmológicos. Este bloqueo se incrementa con el tiempo y la incidencia de complicaciones es baja.

Introduction:Peribulbar anesthesia is currently considered the regional anesthetic technique of choice for various ophthalmic surgical procedures because of its effectiveness and low incidence of complications. Recent techniques have reduced the number of injections from two to one and are undergoing evaluation. Objectives:To measure the efficacy and safety of the caruncular single injection peribulbar technique for various surgical procedures performed at an ophthalmic clinic in Popayan city, Colombia. Methods:Patients undergoing various ophthalmic procedures were included. The anesthetic technique used was based on a caruncular single peribulbar injection. The anesthetic agent used contained 0.5% bupivacaine 2.5 ml, 2% lidocaine 2.5 ml with hyaluroni-dase 7 IU/ml. Patients were evaluated at 10, 15, and 20 min. The motor functionality of the four extraocular muscles and the upper and lower eyelid motor control was measured. Results: 137 patients were included; 54% were females and 77% ASA II. 10 minutes into the evaluation, 92% of the patients achieved an adequate level of anesthesia to proceed with surgery. 36 patients (26.3%) required a booster dose of an additional peribulbar injection. Twenty-two patients (16%) reported mild pain during the anesthetic procedure. There were four cases of chemosis (3%). Conclusions: Using the caruncular single peribulbar injection, most patients achieved an appropriate ocular block for multiple ophthalmic surgical procedures. As time elapses, this block becomes stronger and the incidence of complications is low.

Perturbing pro-survival proteins using quinoxaline derivatives: a structure-activity relationship study.

In HeLa cells the combinatorial knockdown of Bcl-xL and Mcl-1 is sufficient to induce spontaneous apoptosis. Quinoxaline derivatives were screened for the induction of Mcl-1 dependent apoptosis using a cell line without functional Bcl-xL. Quinoxaline urea analog 1 h was able to specifically induce apoptosis in an Mcl-1 dependent manner. We demonstrate that even small changes to 1h results in dramatic loss of activity. In addition, 1 h and ABT-737 synergistically inhibit cell growth and induce apoptosis. Our results also suggest that 1h could have therapeutic potential against ABT-737 refractory cancer.

Active Bax and Bak are functional holins.

The mechanism of Bax/Bak-dependent mitochondrial outer membrane permeabilization (MOMP), a central apoptotic event primarily controlled by the Bcl-2 family proteins, remains not well understood. Here, we express active Bax/Bak in bacteria, the putative origin of mitochondria, and examine their functional similarities to the λ bacteriophage (λ) holin. As critical effectors for bacterial lysis, holin oligomers form membrane lesions, through which endolysin, a muralytic enzyme, escapes the cytoplasm to attack the cell wall at the end of the infection cycle. We found that active Bax/Bak, but not any other Bcl-2 family protein, displays holin behavior, causing bacterial lysis by releasing endolysin in an oligomerization-dependent manner. Strikingly, replacing the holin gene with active alleles of Bax/Bak results in plaque-forming phages. Furthermore, we provide evidence that active Bax produces large membrane holes, the size of which is controlled by structural elements of Bax. Notably, lysis by active Bax is inhibited by Bcl-xL, and the lysis activity of the wild-type Bax is stimulated by a BH3-only protein. Together, these results mechanistically link MOMP to holin-mediated hole formation in the bacterial plasma membrane.

2,3-Substituted quinoxalin-6-amine analogs as antiproliferatives: a structure-activity relationship study.

The quinoxaline core is considered a privileged scaffold as it is found in a variety of biologically relevant molecules. Here we report the synthesis of a quinoxalin-6-amine library, screening against a panel of cancer cell lines and a structure-activity relationship (SAR). This resulted in the identification of a bisfuranylquinoxalineurea analog (7c) that has low micromolar potency against the panel of cancer cell lines. We also show that cells treated with quinoxalineurea 7c results in caspase 3/7 activation, PARP cleavage and Mcl-1 dependent apoptosis.

Perturbation of the Bcl-2 network and an induced Noxa/Bcl-xL interaction trigger mitochondrial dysfunction after DNA damage.

How most apoptotic stimuli trigger mitochondrial dysfunction remains to be resolved. We screened the entire Bcl-2 network for its involvement in DNA damage-induced apoptosis in HeLa cells. Although the anti-apoptotic member Bcl-xL served as a major suppressor, apoptosis initiated only when both Mcl-1 and Bcl-xL were eliminated. The pro-apoptotic members Bak, Bad, Bim, and Noxa were required for apoptosis induced by DNA damaging agents camptothecin and UV. We, therefore, used a His-tagged Bcl-xL expression system to capture the relevant BH3-only proteins that bind to Bcl-xL in response to DNA damage. Surprisingly, unlike Bad and Bim, which bound Bcl-xL constitutively, Noxa became "Mcl-1-free" and interacted with Bcl-xL after DNA damage but not after death receptor engagement. Similar observations were also made in A431 cells. Importantly, this induced interaction caused cytochrome c release and apoptosis and was directly inhibited by Mcl-1, a protein eliminated or inactivated after DNA damage. These results suggest that the loss/inactivation of Mcl-1 in conjunction with an induced Noxa/Bcl-xL interaction may serve as a trigger for mitochondrial dysfunction during DNA damage-induced apoptosis.

Intususcepción ileocólica secundaria a una masa ileal en una mujer de 90 años: reporte de un caso / ileocolic intussusception secundary to an ileal mass in a 90 year old women: a case report

El artículo reporta un caso de intususcepción en una mujer de 90 años de edad. El caso es relevante, pues si bien la invaginación sin patología asociada es infrecuente, lo es aún más la intususcepción con patología subyacente. En muchas ocasiones el imagenólogo define una posibilidad frente a un caso de difícil diagnóstico clínico. El reporte representa para el radiólogo no sólo un reto en la ubicación de la intususcepción, sino en la diferenciación de la enfermedad subyacente y de la masa que la suele acompañar. Pese a que las imágenes son características, es importante ubicar la lesión en la anatomía del intestino. Todo lo anterior con el fin de definir o descartar una posible cirugía innecesaria.

Changes in reproductive physiology of lactating dairy cows due to elevated steroid metabolism.

This manuscript focuses on potential changes in reproductive physiology that occur due to high milk production in lactating dairy cows. Four reproductive measures are discussed: interval to first ovulation, conception rate, duration of estrus, and multiple ovulation rate. The last two responses have now been closely linked to level of milk production. In contrast, time to first ovulation does not appear to be associated with level of milk production, and the association of conception rate with level of milk production is still controversial. In an attempt to explain some of the changes in reproductive physiology caused by high milk production a model of elevated steroid metabolism in lactating dairy cows is presented. Although many aspects of this model remain to be tested, a central role for elevated steroid metabolism in lactation-induced reproductive changes seems likely.

Reproductive hormones and follicular growth during development of one or multiple dominant follicles in cattle.

The mechanisms regulating ovulation rate under natural conditions are not yet defined, particularly for monovular species. In the present study, we evaluated ovarian structures (every 12 h by ultrasonography) and circulating hormones (every 6 h) to determine the differences between cows that developed one (single dominant; n = 16), two (double dominant; n = 8), or three (triple dominant; n = 3) dominant follicles. The four largest follicles were tracked retrospectively, and the data were normalized to the time of expected follicular deviation (F1 >/= 8.5 mm; hour 0). Follicular dynamics from emergence to deviation were similar, whereas after deviation, expected subordinate follicles continued to grow at a rate similar to the dominant follicle. Triple dominants had greater FSH than double dominants (hour -24 to hour -12) and single dominants (hour -42 to hour -6), and double dominants had greater FSH than single dominants (hour -24 to hour -12). Increased circulating estradiol but lower inhibin were observed in cows that developed multiple follicles. In addition, double dominants had greater LH than single dominants (hour -42 to hour -24 and hour -6 to hour 0) and lower progesterone than single dominants (hour -12 and hour -6). Luteal volume was similar between groups, but milk production was greater for codominant than for single-dominant cows. Thus, selection of multiple dominant follicles during high milk production is related to a transient increase in circulating FSH and LH during the 24 h before follicular selection, producing continued postdeviation growth of follicles that ordinarily would have regressed. Increased FSH and LH probably result from decreased circulating inhibin and progesterone in cows that develop codominant follicles.