Detección de Chlamydia spp. en leucocitos de sangre periférica de ovinos sin aborto previo / Detection of Chlamydia spp. In Peripheral Blood Leukocytes in Ewes without any Previous Abortion

Resumen El aborto enzootico ovino es una enfermedad causada por Chlamydia abortus. Es considerada una zoonosis y una de las principales causas de pérdidas económicas en estas explotaciones. Este trabajo se enfocó en utilizar el cultivo de leucocitos de animales sin signos de abortos y la detección de anticuerpos para determinar la posible presencia de C. abortus en explotaciones de traspatio. Se obtuvieron 42 muestras de sangre periférica de ovejas de diferentes poblaciones. La detección de Chlamydia abortus se realizó mediante la tinción de Giemsa y la técnica de PCR. La detección de anticuerpos anti-C. abortus se dio mediante una técnica de ELISA comercial. Los resultados mostraron 21 muestras positivas mediante la técnica de PCR, de las cuales solo 10 fueron positivas mediante la técnica de Giemsa, mientras que 22 sueros mostraron anticuerpos anti-C. abortus. En este estudio el 38,1 % de las muestras fueron positivas a la infección por C. abortus, como se confirmó mediante PCR y serología. En conclusión, los leucocitos de sangre periférica pueden ser útiles para detectar una infección por Chlamydia spp. en explotaciones sin historial de abortos, con lo que se puede conocer la prevalencia real del aborto enzootico ovino en México.

Abstract The Ewes Enzootic Abortion is a disease caused byChlamydia abortus. It is deemed a zoonosis and one of the leading causes of financial losses in this type of business. This article focuses on using the culture of leukocytes from animals without any abortion symptoms and antibody detection to determine the potential presence ofC. abortusin backyard exploitations. Forty-two samples of peripheral blood were obtained from ewes in different populations. The detection ofChlamydia abortuswas carried out by using the Giemsa dye and PCR technique. Anti-C. Abortusantibody detection was performed through a commercial ELISA technique. Results showed 21 positive samples using the PCR, and only ten were positive according to the Giemsa dye, while 22 serum samples showed anti-C. abortusantibody. In this study, 38.1% of the samples were positive for theC. abortusinfection, as verified with the PCR and serology. In conclusion, peripheral blood leukocytes can be helpful to detect an infection caused byChlamydiaspp. Animal exploitation without any previous abortion allows knowing the real prevalence of ewes' enzootic abortion in Mexico.

Ultraestructura del ovocito maduro y en reabsorción de Spondylus limbatus G. B. Sowerby II, 1847 (= S. calcifer ) (Bivalvia: Spondylidae) / Ultrastructure of the mature and in resorption oocytes of Spondylus limbatus G. B. Sowerby II, 1847 (= S. calcifer ) (Bivalvia: Spondylidae)

Spondylus limbatus es una especie bajo protección especial en México, de la que existe poca información biológica y nada sobre estudios histológicos o de ultraestructura del ovario. El objetivo de esta investigación fue caracterizar la morfología ultraestructural de los gametos femeninos maduros y en degeneración. La gónada femenina de S. limbatus en estado de madurez presentó ovocitos postvitelogénicos de 60-70 µm de diámetro, que presentan el aspecto característico de células metabólicamente activas y altamente sintetizadoras. La membrana citoplasmática posee especializaciones destinadas a aumentar la superficie de absorción de la célula, las microvellosidades; el citoplasma presenta numerosos sistemas membranosos relacionados con la síntesis de material de reserva y secreción; y el patrón de organización nuclear altamente lobulado, y por consiguiente con una gran superficie que asegura el intercambio núcleo-citoplasma, se incorpora de forma estructural al proceso de vitelogénesis. Finalmente, se describen los cambios ultraestructurales resultantes de la lisis de los ovocitos: colapso de las membranas nuclear y citoplásmica, y presencia de células hemocíticas macrófagas.

Spondylus limbatus is a species under special protection in Mexico, of which there is little or no information in the literature of biological, histological or ultrastructural studies of the ovary. The objective of this research was to characterize the ultrastructural morphology of mature and degenerating female gametes. The female gonad of S. limbatus in mature state presented post-vitellogenic oocytes 60-70 µm in diameter, which have characteristics of metabolically active and highly synthesizing cells. The cytoplasmic membrane has specializations designed to increase the absorption surface of the cell, the microvilli; the cytoplasm presents numerous membranous systems related to synthesis of reserve and secretion material as well as the highly lobed nuclear organization pattern; a large surface that ensures core-cytoplasm exchange, is structurally incorporated into the vitellogenesis process. Finally, ultrastructural changes resulting from the lysis of the oocytes are described: collapse of nuclear and cytoplasmic membranes, and the presence of macrophage hemocytic cells.

Pathogenic determinants of clinical Klebsiella pneumoniae strains associated with their persistence in the hospital environment

Background: Klebsiella pneumoniae is considered an opportunistic pathogen associated with nosocomial infections occurring mainly in pediatric patients, such as premature infants placed in intensive care units. The aim of this study was to characterize K. pneumoniae strains isolated from different clinical sources based on their resistance to antibiotics and the presence of virulence factors associated with their persistence in the hospital environment. Methods: Fifty clinical strains of K. pneumoniae isolated from urine, blood, catheters, and cerebrospinal fluid sources were characterized. Susceptibility testing of antibiotics was performed by the Kirby-Bauer method (Clinical Laboratory Standards Institute, 2010). The ability to form a biofilm was determined by the 96-well microplate method. Capsule and fimbrial structures were visualized by transmission electron microscopy (TEM). Adherence was evaluated on A549 and HT29 cells. Assessment for the presence and expression of the ecpA, fimH, and mrkA genes was performed by PCR and RT-PCR. Results: Clinical strains of K. pneumoniae were isolated from 48% of urine, 24% of blood, 18% of catheters, and 10% of cerebrospinal fluid. Ninety-two percent of the strains showed resistance to cefpodoxime, whereas few strains showed resistance to imipenem and meropenem (4 and 2%, respectively). The extended spectrum-type beta-lactamase (ESBL) phenotype was identified in 97% of the strains positive for resistance to third-generation cephalosporins. In addition, 88% of the strains were multidrug resistant. All strains were able to form biofilms. Capsule and fimbirial structures were visualized by TEM. Based on our adhesion assays, the A549 cell line was more permissive to K. pneumoniae strains than the HT-29 cell line. K. pneumoniae strains amplified and expressed ecpA (100/70%), fimH (98/2%), and mrkA (84/48%) genes, respectively. Conclusion: The K. pneumoniae strains exhibited features that allowed them to survive in the hospital environment (formation of biofilm) and resist antimicrobial therapy (multidrug resistant MDR strains). These strains also possessed a capsule, adhesive properties, and expression of genes encoding colonization factors that favor the selection and persistence of these strains in hospitals.

Análisis in silico de proteínas potencialmente involucradas en la biogénesis de fimbrias en Helicobacter pylori / In silico analysis of proteins potentially involved in fimbrial biogenesis in Helicobacter pylori

Introducción. La colonización e infección crónica por Helicobacter pylori es el factor de mayor contribución al desarrollo de cáncer gástrico. Se ha descrito un gran repertorio de adhesinas que contribuyen a la adaptación específica de la bacteria al nicho gástrico y, para H. pylori , al igual que en otras bacterias patógenas, la formación de biopelícula es fundamental en la supervivencia a ambientes no favorables. Las fimbrias o pili tipo IV son responsables de la adhesión de diversas bacterias patógenas ( Escherichia coli , Pseudomonas aeruginosa y Vibrio cholerae) a distintas superficies. El objetivo de este trabajo fue identificar y analizar genes que pudieran codificar para proteínas involucradas en la biogénesis de fimbrias en H. pylori y caracterizar su expresión durante la formación de biopelícula. Métodos. Se emplearon herramientas bioinformáticas y moleculares, tales como la base de datos del NCBI para la búsqueda de secuencias de proteínas relacionadas con la biogénesis de fimbrias, así como la herramienta de PSI BLAST. Los alineamientos múltiples se realizaron con el programa T-COFFEE y HMMER. La predicción de las estructuras secundarias se realizó con ANTHEPROT y las estructuras terciarias se predijeron con el programa I-TASSER. Resultados. Se identificaron dos homólogos, jhp0257 y HP0272, de la proteína PilN de Campylobacter rectus y Xilella fastidiosa , la cual es parte de la maquinaria del ensamble de la fimbria tipo IV. Asimismo, las proteínas jhp0887 y HP0953 presentaron homología a nivel del péptido señal de PilA de P. aeruginosa , y la proteína HP0953 se sobreexpresó durante la formación de la biopelícula. Conclusiones. H. pylori posee proteínas homólogas a las proteínas de familias fimbriales, específicamente PilN y PilA, que ensamblan fimbria tipo IV en otras bacterias. Esta última tiene un nivel de expresión mayor durante la etapa inicial del proceso de formación de biopelícula.

Background. Colonization and chronic infection with Helicobacter pylori is the major contributing factor to the development of gastric cancer. A large repertoire of adhesins has been described that contribute to the adaptation of bacteria to a specific gastric niche. As in other pathogenic bacteria, H. pylori biofilm formation is central to survival on unfavorable environments. Type IV pili or fimbriae are responsible for the adhesion of many pathogenic bacteria (e.g., Escherichia coli, Pseudomonas aeruginosa and Vibrio cholerae ) to various surfaces. The aim of this study was to identify and analyze genes that might encode proteins involved in the biogenesis of fimbriae on H. pylori and characterize their expression during biofilm formation. Methods. PSI BLAST, bioinformatics and molecular tools were used as well as the NCBI database search for sequences related to protein biogenesis of fimbriae. Multiple alignments were performed using the HMMer and T-COFFEE programs. The secondary structure prediction was performed with ANTHEPROT and the tertiary structures were predicted with the I-Tasser. Results. We identified two counterparts-jhp0257 and HP0272-from protein of Campylobacter rectus and PilN Xilella fastidiosa , which is part of the machinery of assembly type IV fimbria. Similarly, proteins jhp0887 and HP0953 show homology from peptide PilA level of P. aeruginosa , and the HP0953 protein is overexpressed during the formation of the biofilm. Conclusions. H. pylori possesses proteins homologous to fimbrial protein families, specifically PilN and PilA, which join type IV fimbriae in other bacteria. The latter has a higher expression level during the initial stage of the formation of biofilm.

Descripción histológica de la cámara incubatriz de moina hutchinsoni (Brem, 1937) / Histological description of the brood-chamber of moina hutchinsoni (Brem, 1937)

Los cladóceros son partenogenéticos por lo que la mayor parte del año, las poblaciones consisten enteramente de hembras que se reproducen asexualmente, en ellas el ovario se comunica por medio de un oviducto, con la cámara incubatriz, la cual se localiza en el margen interno posterior del caparazón, cerca del corazón y antes del intestino. Los huevos provenientes de los oviductos se depositan en la cámara y se incuban hasta terminar el desarrollo embrionario. Se considera que Moina presenta un desarrollo postembrionario directo, porque los organismos juveniles o neonatos, salen completamente formados e independientes durante la muda. En la reproducción asexual la cámara contiene a las diferentes etapas del desarrollo embrionario llamadas; ovocito, huevo y embrión. En la etapa sexual o gamogenética la cámara contiene un efipio con dos huevos. La cámara incubatriz histológicamente esta conformada por un epitelio plano simple, que descansa sobre una membrana basal evidente, la cual se continúa con el tejido conjuntivo laxo, después del cual se encuentra el caparazón. En el interior de la cámara se identificaron los ovocitos, huevos y algunas etapas del desarrollo embrionario, en cortes semifinos y finos por microscopia óptica y de transmisión, respectivamente.

Cladocerans are parthenogenetic during the greater part of the year. Populations consist entirely of females that reproduce asexually; in these females the ovary communicates through an oviduct with the brood chamber which is located at the rear inner shell border, near the heart and before the intestine. The eggs originating from the oviducts are deposited in the brood chamber and are incubated until the end of embryonic development. Moina is considered post embryonic development because juvenile or neonate organisms emerge fully formed and independent during moulting. In asexual reproduction the brood chamber contains the various stages of embryonic development denominated, oocyte, egg and embryo. At this stage the sexual gamogenetic contains an ephippium with two eggs. The brood chamber is formed histologically by a simple, plane epithelium that rests on a clear basal membrane continuous with the loose connective tissue, after which the shell can be located. Inside the brood chamber oocytes, eggs and some stages of embryonic development are identified, as well as fine semi optical and transmission microscopy cuts respectively.