First report of caprine abortions due to Chlamydia abortus in Argentina.

Infectious abortions of goats in Argentina are mainly associated with brucellosis and toxoplasmosis. In this paper, we describe an abortion outbreak in goats caused by Chlamydia abortus. Seventy out of 400 goats aborted. Placental smears stained with modified Ziehl-Neelsen stain showed many chlamydia-like bodies within trophoblasts. One stillborn fetus was necropsied and the placenta was examined. No gross lesions were seen in the fetus, but the inter-cotyledonary areas of the placenta were thickened and covered by fibrino-suppurative exudate. The most consistent microscopic finding was found in the placenta and consisted of fibrinoid necrotic vasculitis, with mixed inflammatory infiltration in the tunica media. Immunohistochemistry of the placenta was positive for Chlamydia spp. The results of polymerase chain reaction targeting 23S rRNA gene performed on placenta were positive for Chlamydia spp. An analysis of 417 amplified nucleotide sequences revealed 99% identity to those of C. abortus pm225 (GenBank AJ005617) and pm112 (GenBank AJ005613) isolates. To the best of our knowledge, this is the first report of abortion associated with C. abortus in Argentina

Distribución y aspectos poblacionales del lobo Ibérico en la provincia de Ourense

Para determinar el área de distribución del lobo en la provincia de Ourense (noroeste de España) se realizaron prospecciones de campo consistentes en itinerarios de muestreo para localizar indicios y se enviaron cuestionarios a los Agentes del Servicio de Conservación de la Naturaleza de la Xunta de Galicia y a las Sociedades de Cazadores del área de estudio. Los resultados se registraron sobre cuadrículas U.T.M. 10x10 Km. Se constata presencia del lobo en el 79,2% de las cuadrículas estudiadas, estimando el área de distribución en la provincia en 6.400 km2. Para la localización de los grupos familiares inicialmente se prospectó el territorio a nivel de cuadrículas U.T.M. 10x10 Km y en función de los resultados y la información recibida se eligieron determinadas zonas para realizar estaciones de escucha y espera. Determinamos durante el periodo de estudio la existencia de 25 grupos familiares. La densidad estimada en la provincia es de 2,10 – 3,28 lobos /100 km2. De los casos de mortalidad conocidos entre enero de 1999 y abril de 2002 (n=37), un elevado porcentaje se debió a atropellos (70,27%) principalmente en dos vías concretas y un 8,10% correspondió a envenenamientos. Actualmente la densidad del lobo en Ourense es superior a la obtenida en provincias limítrofes, pero amenazas de origen antrópico tanto directas (veneno, persecución ilegal...) como indirectas (incendios forestales, grandes infraestructuras viales, parques eólicos...) pueden estar condicionando la existencia de grupos familiares e incluso la presencia de la especie en determinadas zonas

Clostridium perfringens epsilon toxin induces blood brain barrier permeability via caveolae-dependent transcytosis and requires expression of MAL.

Clostridium perfringens epsilon toxin (ETX) is responsible for causing the economically devastating disease, enterotoxaemia, in livestock. It is well accepted that ETX causes blood brain barrier (BBB) permeability, however the mechanisms involved in this process are not well understood. Using in vivo and in vitro methods, we determined that ETX causes BBB permeability in mice by increasing caveolae-dependent transcytosis in brain endothelial cells. When mice are intravenously injected with ETX, robust ETX binding is observed in the microvasculature of the central nervous system (CNS) with limited to no binding observed in the vasculature of peripheral organs, indicating that ETX specifically targets CNS endothelial cells. ETX binding to CNS microvasculature is dependent on MAL expression, as ETX binding to CNS microvasculature of MAL-deficient mice was not detected. ETX treatment also induces extravasation of molecular tracers including 376Da fluorescein salt, 60kDA serum albumin, 70kDa dextran, and 155kDA IgG. Importantly, ETX-induced BBB permeability requires expression of both MAL and caveolin-1, as mice deficient in MAL or caveolin-1 did not exhibit ETX-induced BBB permeability. Examination of primary murine brain endothelial cells revealed an increase in caveolae in ETX-treated cells, resulting in dynamin and lipid raft-dependent vacuolation without cell death. ETX-treatment also results in a rapid loss of EEA1 positive early endosomes and accumulation of large, RAB7-positive late endosomes and multivesicular bodies. Based on these results, we hypothesize that ETX binds to MAL on the apical surface of brain endothelial cells, causing recruitment of caveolin-1, triggering caveolae formation and internalization. Internalized caveolae fuse with early endosomes which traffic to late endosomes and multivesicular bodies. We believe that these multivesicular bodies fuse basally, releasing their contents into the brain parenchyma

Distribución y aspectos poblacionales del lobo Ibérico (Canis lupus signatus) en las provincias de Pontevedra y A Coruña (Galicia)

Para determinar el área de distribución del lobo en las provincias de Pontevedra y A Coruña se realizaron itinerarios de muestreo y se enviaron cuestionarios a los Agentes de Medio Ambiente de la Xunta de Galicia y a las Sociedades de Cazadores del área de estudio. Los resultados se representaron en cuadrículas U.T.M. 10x10 Km. Se ha confirmado la presencia del lobo en el 67,05% de las cuadrículas del área de estudio. La especie se distribuye de forma continua en ambas provincias, donde estimamos un área de distribución de 6.400 km2 en la provincia de A Coruña y 3.200 km2 en la de Pontevedra, ocupando principalmente el área montañosa de la Dorsal Gallega y el occidente de A Coruña; esta última zona está conectada con el resto del área de distribución del lobo en Galicia a través del norte de la provincia de Pontevedra. Para la localización de las manadas se muestreó el territorio a nivel de cuadrícula U.T.M. 10x10 Km, y en las zonas donde se detectó una mayor concentración de indicios, se realizaron estaciones de escucha y espera con el fin de localizar la presencia de manadas (cachorros y adultos). Se determinó la presencia de 29 manadas (14 en la provincia de A Coruña y 15 en Pontevedra), de las cuales se confirmó la reproducción del 52%. Se ha estimado un tamaño poblacional para la provincia de A Coruña entre 87-129 lobos (1,36-2,02 lobos/100 Km2) y entre 95-140 para la provincia de Pontevedra (2,97-4,38 lobos/100 Km2). Las zonas de mayor densidad se localizaron en la Dorsal Gallega. Se han localizado algunas manadas en áreas muy humanizadas, lo que podría explicar que la principal causa de mortalidad conocida entre enero de 2000 y octubre de 2003 (n=29) sea el atropello (n=21)

Sorological evaluation of commercial vaccines against enterotoxemia in goats.

Enterotoxemia in sheep and in goats is caused by the effects of the epsilon toxin of C/ostridium perfringens type D, being considered the main infectious cause of mortality in those animal species. The main prophylactic measures include adequate nutritional management and vaccination of ali animais using vaccines of high immunogenic power. Six commercial vaccines containing in its formulation the epsilon toxoid of C. perfringens type D were sorogically evaluated. Eighty four female goat kids, whose mothers had no previous vaccination history against clostridioses were used. They were divided into six groups of 14 animais each. The animais of the control group didn't receive any vaccine dose and the animais from the groups 1 to 5 received two vaccine doses, The first vaccine dose was applied at 45 days of life (day zero) and the second dose at 75 days (30 days after the first dose). Blood samples were collected from the goat kids at the days zero, 30, 60, 90, 120 and 150 after the beginning of the experiment, in order to evaluate the immunologic response. The Indirect ELlSA technique was used for the quantification of the antibodies against epsilon toxin in the samples of blood serum of the animais. In day zero, no animal presented titre considered protector. The largest number of animais considered protected was found at day 60, in response to the two initial doses of the vaccine (days O and 30, first and second doses, respectively). Only tive animaIs which received the vaccine 1 and one animal which received the vaccine 3 stayed wilh titres of antibodies considered up to 150 days after the first vaccine dose. Based on the results, it was concluded lhat the evaluated vaccines showed small amount of epsilon toxoid in the commercial formulations, a crucial fact for lhe low efficiency of. the vaccines. For commercial reasons, the vaccines against the clostridioses present versatile formulations, with several toxoid types, used for various animal species, which certainly contributed to reduce their effectiveness in preventing the iIInesses caused by the clostridia or their toxins

Agromorphological characterization and dollar spot fungus susceptibility in accessions of common bent (Agrostis capillaris L.) collected in northern Spain

Abstract Fourteen Agrostis capillaris accessions collected in northern Spain were characterized in a trial with a lowfertilization regime, carried out in two successive years (2004 and 2005). The commercial cultivar 'Golfin' was included in the study as a control. All accessions and the commercial cultivar were evaluated for 11 agromorphological characteristics and susceptibility to dollar spot disease, caused by Sclerotinia homoeocarpa F.T. Bennet. The data obtained were reduced to five principal components that cumulatively explained 85.4% of the total variance. Cluster analysis was useful in identifying four groups of accessions that described 50% of the phenotypic variation among accessions. Cluster 1 consisted of four accessions with the highest resistance to dollar spot, dark green colour, late heading date and high autumn turf quality. Cluster 2 included the two earliest heading accessions, dark green colour, intermediate tolerance to dollar spot and low autumn turf quality. Cluster 3 comprised six accessions with the latest heading date, dark green colour and low tolerance to dollar spot. Cluster 4 consisted of two accessions and the 'Golfin' cultivar, with lightest green colour, low autumn turf quality and the poorest tolerance to dollar spot. The results of this study suggest the potential value of some of these common bent accessions collected in northern Spain for turf and forage improvement. Additional key words: Agrostis tenuis, artificial inoculation, browntop, colonial bentgrass, multivariate analysis, turfgrass. Resumen Caracterización agromorfológica y susceptibilidad al hongo «dollar spot» en accesiones de agrostis común (Agrostis capillaris L.) recogidas en el Norte de España Catorce accesiones de Agrostis capillaris procedentes del norte de España se caracterizaron en un régimen de baja fertilización durante dos años (2004 y 2005) en un diseño de bloques completos al azar con dos repeticiones de 15 plantas por accesión (en total 30 plantas por accesión). El cultivar comercial 'Golfin' se incluyó como control en el estudio. Todas las accesiones y el cultivar 'Golfin' se evaluaron mediante 11 caracteres agromorfológicos y la susceptibilidad a la enfermedad fúngica «dollar spot» (Sclerotinia homoeocarpa F.T. Bennet). Los datos obtenidos se redujeron a cinco componentes principales que explicaron el 85,4% de la varianza total. Mediante una clasificación jerárquica se identificaron cuatro grupos, que describen el 50% de la variación fenotípica entre las accesiones. El grupo 1 incluyó cuatro accesiones con la mayor resistencia al «dollar spot», color verde oscuro, espigado tardío y muy buena calidad de césped en el otoño. El grupo 2 incluyó las dos accesiones más precoces de espigado, tolerancia intermedia al «dollar spot» y mala calidad de césped en el otoño. El grupo 3 comprendió seis accesiones tardías de espigado, color verde oscuro y baja tolerancia al «dollar spot». El grupo 4 incluyó dos accesiones y el cultivar 'Golfin' con color verde más claro, baja calidad otoñal de césped y la menor tolerancia al dollar spot. Los resultados de este estudio muestran el potencial de algunas de las accesiones de agrostis común procedentes del norte de España para uso en mejora de céspedes y forrajes. Palabras clave adicionales: Agrostis tenuis, análisis multivariable, césped, inoculación artificial

Gene-Trap Mutagenesis Identifies Mammalian Genes Contributing to Intoxication by Clostridium perfringens ε-Toxin

The Clostridium perfringens ε-toxin is an extremely potent toxin associated with lethal toxemias in domesticated ruminants and may be toxic to humans. Intoxication results in fluid accumulation in various tissues, most notably in the brain and kidneys. Previous studies suggest that the toxin is a pore-forming toxin, leading to dysregulated ion homeostasis and ultimately cell death. However, mammalian host factors that likely contribute to ε-toxin-induced cytotoxicity are poorly understood. A library of insertional mutant Madin Darby canine kidney (MDCK) cells, which are highly susceptible to the lethal affects of ε-toxin, was used to select clones of cells resistant to ε-toxin-induced cytotoxicity. The genes mutated in 9 surviving resistant cell clones were identified. We focused additional experiments on one of the identified genes as a means of validating the experimental approach. Gene expression microarray analysis revealed that one of the identified genes, hepatitis A virus cellular receptor 1 (HAVCR1, KIM-1, TIM1), is more abundantly expressed in human kidney cell lines than it is expressed in human cells known to be resistant to ε-toxin. One human kidney cell line, ACHN, was found to be sensitive to the toxin and expresses a larger isoform of the HAVCR1 protein than the HAVCR1 protein expressed by other, toxin-resistant human kidney cell lines. RNA interference studies in MDCK and in ACHN cells confirmed that HAVCR1 contributes to ε-toxin-induced cytotoxicity. Additionally, ε-toxin was shown to bind to HAVCR1 in vitro. The results of this study indicate that HAVCR1 and the other genes identified through the use of gene-trap mutagenesis and RNA interference strategies represent important targets for investigation of the process by which ε-toxin induces cell death and new targets for potential therapeutic intervention

Genome Sequencing and Analysis of a Type A Clostridium perfringens Isolate from a Case of Bovine Clostridial Abomasitis

Clostridium perfringens is a common inhabitant of the avian and mammalian gastrointestinal tracts and can behave commensally or pathogenically. Some enteric diseases caused by type A C. perfringens, including bovine clostridial abomasitis, remain poorly understood. To investigate the potential basis of virulence in strains causing this disease, we sequenced the genome of a type A C. perfringens isolate (strain F262) from a case of bovine clostridial abomasitis. The ∼3.34 Mbp chromosome of C. perfringens F262 is predicted to contain 3163 protein-coding genes, 76 tRNA genes, and an integrated plasmid sequence, Cfrag (∼18 kb). In addition, sequences of two complete circular plasmids, pF262C (4.8 kb) and pF262D (9.1 kb), and two incomplete plasmid fragments, pF262A (48.5 kb) and pF262B (50.0 kb), were identified. Comparison of the chromosome sequence of C. perfringens F262 to complete C. perfringens chromosomes, plasmids and phages revealed 261 unique genes. No novel toxin genes related to previously described clostridial toxins were identified: 60% of the 261 unique genes were hypothetical proteins. There was a two base pair deletion in virS, a gene reported to encode the main sensor kinase involved in virulence gene activation. Despite this frameshift mutation, C. perfringens F262 expressed perfringolysin O, alpha-toxin and the beta2-toxin, suggesting that another regulation system might contribute to the pathogenicity of this strain. Two complete plasmids, pF262C (4.8 kb) and pF262D (9.1 kb), unique to this strain of C. perfringens were identified