Neospora caninum antibodies in wild carnivores from Spain

Serum samples from 251 wild carnivores from different regions of Spain were tested for antibodies to Neospora caninum by the commercial competitive screening enzyme linked immunosorbent assay (c-ELISA) and confirmed by Neospora agglutination test (NAT) and/or by indirect fluorescent antibody test (IFAT). Samples with antibodies detected by at least two serological tests were considered seropositive. Antibodies to N. caninum were found in 3.2% of 95 red foxes (Vulpes vulpes); in 21.4% of 28 wolves (Canis lupus); in 12.0% of 25 Iberian lynx (Lynx pardinus); in 16.7% of 6 European wildcats (Felis silvestris); in 6.4% of 31 Eurasian badgers (Meles meles); in 21.4% of 14 stone martens (Martes foina); in 66.7% of 3 pine martens (M. martes) and in 50% of 2 polecats (Mustela putorius). Antibodies to N. caninum in common genets (Genetta genetta) and Egyptian mongooses (Herpestes ichneumon) were only observed by c-ELISA but were not confirmed by IFAT and/or NAT. No antibodies were detected in 5 Eurasian otters (Lutra lutra) by any technique. Statistically significant differences were observed among species and among geographical areas. The highest seroprevalence of N. caninum infection was observed in the Cantabric Coastal region characterized by high humidity. To our knowledge, this is the first report of antibodies to N. caninum in free ranging wild carnivores, other than wild canids, in Europe. The existence of a possible sylvatic cycle could have important implications in both sylvatic and domestic cycles since they might influence the prevalence of infection in cattle farms in those areas.This study is a contribution to GC-05-006, ICS, JCCM, and agreements between IREC and Principado de Asturias and Ministerio de Agricultura and OAPN, Spain. This study received partially support from the Spanish CICYT, grants AGL2004-06103, AGL2005-07401 and AGL2007-65521. This study was also partially included in the “Programa de Actuaciones para la Conservación del Lince en Andalucía II”. Consejería del Medio Ambiente, Junta de Andalucía. R Sobrino was recipient of a Ph.D. grant from Junta de Comunidades de Castilla La Mancha (JCCM).Peer reviewe

Genomic profiling of primary and recurrent Adult Granulosa Cell Tumors of the Ovary

Adult-type granulosa cell tumor (aGCT) is a rare malignant ovarian sex cord-stromal tumor, harboring recurrent FOXL2 c.C402G/p.C134W hotspot mutations in 97% of cases. These tumors are considered to have a favorable prognosis, however aGCTs have a tendency for local spread and late recurrences, which are associated with poor survival rates. We sought to determine the genetic alterations associated with aGCT disease progression. We subjected primary non-recurrent aGCTs (n = 7), primary aGCTs that subsequently recurred (n = 9) and their matched recurrences (n = 9), and aGCT recurrences without matched primary tumors (n = 10) to targeted massively parallel sequencing of ≥410 cancer-related genes. In addition, three primary non-recurrent aGCTs and nine aGCT recurrences were subjected to FOXL2 and TERT promoter Sanger sequencing analysis. All aGCTs harbored the FOXL2 C134W hotspot mutation. TERT promoter mutations were found to be significantly more frequent in recurrent (18/28, 64%) than primary aGCTs (5/19, 26%, p = 0.017). In addition, mutations affecting TP53, MED12, and TET2 were restricted to aGCT recurrences. Pathway annotation of altered genes demonstrated that aGCT recurrences displayed an enrichment for genetic alterations affecting cell cycle pathway-related genes. Analysis of paired primary and recurrent aGCTs revealed that TERT promoter mutations were either present in both primary tumors and matched recurrences or were restricted to the recurrence and absent in the respective primary aGCT. Clonal composition analysis of these paired samples further revealed that aGCTs display intra-tumor genetic heterogeneity and harbor multiple clones at diagnosis and relapse. We observed that in a subset of cases, recurrences acquired additional genetic alterations not present in primary aGCTs, including TERT, MED12, and TP53 mutations and CDKN2A/B homozygous deletions. Albeit harboring relatively simple genomes, our data provide evidence to suggest that aGCTs are genetically heterogeneous tumors and that TERT promoter mutations and/or genetic alterations affecting other cell cycle-related genes may be associated with disease progression and recurrences

Seroprevalence of Neospora caninum in non-carnivorous wildlife from Spain

Serum samples from 1034 non-carnivorous wildlife from Spain were tested for antibodies to Neospora caninum by competitive screening enzyme linked immunosorbent assay (ELISA) and confirmed by an indirect fluorescent antibody test (IFAT). High agreement was observed between results in both techniques (kappa value higher than 0.9). Prevalences of N. caninum antibodies positive by both techniques were 11.8% of 237 red deer (Cervus elaphus), 7.7% of 13 barbary sheep (Ammotragus lervia), 6.1% of 33 roe deer (Capreolus capreolus) and 0.3% of 298 wild boar (Sus scrofa). In one of 53 hares (Lepus granatensis), antibodies were found in the ELISA but could not be confirmed by IFAT due to lack of sample. Antibodies to N. caninum were not found in any of 251 wild rabbits (Oryctolagus cuniculus), 79 fallow deer (Dama dama), 27 mouflon (Ovis ammon), 40 chamois (Rupicapra pyrenaica) and three Spanish ibex (Capra pyrenaica). Statistically significant differences were observed between N. caninum seroprevalence in red deer and management of hunting estates (open versus fenced) with higher prevalence in fenced estates, and among sampling sites. Seroprevalence was particularly high in some areas (MO estate in South-Central Spain or some estates of Catalonia, North-East Spain), while no contact with N. caninum was observed in others. Results indicate that in certain areas of Spain, N. caninum is present in wildlife, especially in red deer. These results have important implications in both sylvatic cycles and may influence the prevalence of infection in cattle farms in those areas. To our knowledge, this is the first report of antibodies to N. caninum in wildlife from Spain and the first report of N. caninum antibodies in barbary sheep and wild boar.This is a contribution to the agreements between Yolanda Fierro and UCLM, CSIC, OAPN and SDGSA, Ministerio de Agricultura. Fran Ruiz-Fons received financial support through a grant from the Ministerio de Educación y Ciencia. This study received support from the Spanish CICYT, grant AGL2004-06103-C02-01/GAN and AGL2005-07401-CO3-01-GAN, Ministerio de Ciencia y Tecnología and FEDER.Peer reviewe

Aspectos epidemiológicos de la neosporosis bovina en el Nordeste español. Una perspectiva clínica

El parásito protozoo intracelular Neospora caninum ha sido descrito en un amplio rango de especies animales y se considera una de las causas más importantes de aborto en ganado vacuno en todo el mundo. En España, la seroprevalencia individual en las explotaciones y la tasa de infección en fetos abortados es elevada. Revisamos aquí aspectos epidemiológicos de la neosporosis en explotaciones de ganado vacuno lechero de alta producción en el Nordeste español. Es interesante resaltar que independientemente de la seroprevalencia de N. caninum a nivel de explotación, la titulación de los anticuerpos frente el parásito es un buen indicador del riesgo de aborto, que la seropositividad frente N. caninum es muy estable a lo largo del tiempo, de manera que las vacas crónicamente infectadas presentan un alta tasa de abortos repetitivos, y que la infección por N. caninum no afecta a la fertilidad de las vacas infectadas ni compromete la gestación durante el primer trimestre de la misma.Estos trabajos han sido subvencionados por los proyectos CICYT: AGL2000-0904, AGL2004-06103 and AGL2007-65521.Publishe

Managing gestation in cattle

Once a cow becomes pregnant, the effect of pregnancy loss on its reproductive cycle is a topic of great interest for dairy herds. This paper reviews the factors of a non-infectious nature that affect the pregnancy maintenance during the late embryonic/early fetal period. Some clinical suggestions on bovine neosporosis and coxiellosis are also highlighted

Managing gestation in cattle

Once a cow becomes pregnant, the effect of pregnancy loss on its reproductive cycle is a topic of great interest for dairy herds. This paper reviews the factors of a non-infectious nature that affect the pregnancy maintenance during the late embryonic/early fetal period. Some clinical suggestions on bovine neosporosis and coxiellosis are also highlighted

Relationships between peripheral maternal immune status during the peripartum period and postpartum reproductive disorders

Pregnancy and peripartum period are characterised by transient changes in the maternal immune system, noticeable at the peripheral level. The aim of the study was to determinate the effect of placental retention and metritis on peripheral white blood cells in 91 pregnant dairy cows from day 200 of pregnancy to 30 postpartum. Blood samples were automatically analysed with HemaVet® Multispecies Haematology system and total and differential leukocyte counts were recorded. Data was statistically analysed using GLM repeated measures ANOVA to test the effect of reproductive disorders on peripheral white blood cells. Cows with placental retention had significantly lower leukocyte, neutrophil and lymphocyte counts (P<0.05), and cows with metritis had lower neutrophil and eosinophil counts (P<0.01) than cows not suffering reproductive disorders during the peripartum period. Day of pregnancy and parturition also affected peripheral lymphocyte and monocyte counts (P<0.01). Maternal immune dysfunction during the peripartum period may predispose to reproductive disorders postpartu

No detectable precolostral antibody response in calves born from cows with cotyledons positive for Coxiella burnetii by quantitative PCR

Samples from 45 dams (milk/colostrum, faeces, vaginal fluid and blood on days 171–177 of gestation and at parturition, and cotyledons at parturition) and their calves (blood collected before colostrum intake and weekly until days 29–35) were analysed to examine the vertical transmission of Coxiella burnetii and links between shedding and seropositivity. All calves were born C. burnetii seronegative. Only those born to seropositive dams seroconverted following colostrum intake. Logistic regression analyses indicated that the likelihood of dam seropositivity was 21 and 4.85 times higher for multiparous than for primiparous (65.6% vs. 8.3%, P = 0.006) and for prepartum shedding cows (75% vs. 38.2%, P = 0.03) compared to the remaining animals, respectively. In conclusion, the results of this study indicate no detectable precolostral antibody response in calves born from dams with cotyledons positive for C. burnetii by qPCR. In order to analyse the possibility of persistent infection due to immunotolerance to an early in utero infection, further studies will need to test for C. burnetii DNA. In addition, in the present study multiparous cows showed a significantly higher seroprevalence than primiparous cows and heifers, colostral antibodies were efficiently transferred to newborn calves, and there was a link between bacterial shedding on days 171–177 of gestation and Coxiella seropositivity of the dam

Plasma progesterone concentration is not reduced in pregnant non-aborting heifers experimentally infected with neospora caninum

El feto mamífero es antigénicamente un cuerpo extraño para la madre y para sobrevivir se producen una serie de interacciones de inmunosupresión entre la madre y el mismo. La hormona clave que provoca inmunotolerancia fetal por parte de la madre es la progesterona (Hansen et al. 1986), produciendo un cambio hacia la respuesta inmune tipo Th2 y evitando así el aborto que produciría una excesiva respuesta Th1 (Roberston, 2000). El efecto regulador de la progesterona es ejercido directa o indirectamente a través de mediadores secundarios como las serpinas uterinas, conocidas como SERPINA14, que se encuentran presentes en el endometrio, fluidos fetales y el ovario (Padua y Hansen, 2010). Neospora caninum es un protozoo intracelular obligado considerado la causa más importante de aborto en vacuno lechero a nivel mundial. En las granjas lecheras la ruta de infección más frecuente es la transplacentaria, donde las madres infectadas naturalmente transmiten el parasito a los fetos durante la gestación (Almería y López-Gatius, 2013). La mayoría de terneros nacen clínicamente normales, aunque aproximadamente un 95% están infectados por N. caninum. El aborto o la infección congénita ocurren cuando el parásito atraviesa la barrera placentaria e infecta al feto, provocando el aborto principalmente hacia los 5-7 meses de gestación (Almería y López-Gatius, 2013). Las razones por las cuales algunas vacas infectadas abortan y otras no, siguen siendo desconocidas. Es bien conocido que la infección por N. caninum modifica los patrones hormonales durante la gestación en vacuno lechero. Por ejemplo, la seropositividad frente a este parásito ha sido asociada con incremento de prolactina y progesterona plasmática (Garcia-Ispierto et al. 2010, 2013) y reducción de glicoprotefnas asociadas a gestación (PAGs) (Garcia-Ispierto et al., 2015). También, se ha observado una correlación negativa entre la expresión génica de SERP/NA 14 en la placenta y la respuesta adaptativa humoral y celular en la placenta y linfonodos uterinos de vacas infectadas experimentalmente con N. caninum (Serrano-Pérez et al., 2016, 2017). El presente estudio se realizó durante el segundo trimestre de gestación en terneras gestantes experimentalmente infectadas con N. caninum el dla 11 O de gestación. Se seleccionó este momento de infección debido a que es cuando la mayoría de abortos tienen lugar en animales infectados en condiciones de campo. Debido al tropismo del parásito por el folículo ovárico (Silva et al., 2012) y al aumento de progesterona detectado en animales seropositivos a N. caninum, el objetivo de este estudio fue determinar si las novillas infectadas experimentalmente mostraban cambios en Ja concentración plasmática de progesterona tras la infección. Un segundo objetivo fue correlacionar la expresión génica de SERPINA 14 en el cuerpo lúteo con la concentración plasmática de progesterona.Neospora caninum is a majar cause of abortion in cattle worldwide. However,immune-endocrine interactions during pregnancy in Neospora-infected cows remainunknown. This study examines plasma progesterone concentration in dairy heifers experimentally infected with N. caninum that did not abort, and also assessed possible interrelations between expression patterns of SERPINA 14 in corpus luteum tissues and plasma progesterone concentrations. The study population was constituted of tour noninfected controls and three heifers experimentally infected wíth N. caninum on Day 11 O of pregnancy with live foetuses at euthanasia. Both groups were euthanized on Day 152 of gestation. At euthanasia, blood samples were collected to determine progesterone concentrations. Samples of corpus luteum tissues (CL) were collected to determine gene expression of SERPINA14. No relationships were detected between infection and plasma progesterone concentration (12.5 ± 1.5 ng/ml in controls vs. 12.8 ± 3.7 ng/ml in infected dams). There was not significan! correlation between plasma progesterone concentration and SERPINA14 expression in CL (r: 0.607, P=0.148). In conclusion, N. caninum had no impacts on luteal funclion, at least in terms of repercussions on plasma progesterone concentrations.Este estudio fue financiado por MINECO (AGL2012-39830-C02-01/02) y fondos FEDER. R. Mur-Novales disfruta de una beca FPI (BES-2013-063215).Publishe

Un nou model per estudiar l'avortament en les granges de boví infectades per Neospora caninum

Investigadors de la UAB i de la Universitat de Lleida han observat que la infecció en laboratori pel paràsit N. caninum en vaques gestants que es troben en el segon trimestre de gestació és capaç de causar avortament, imitant el que passa a les condicions de granja. L'estudi demostra que és un model adequat per estudiar i tractar d'entendre la patogènia dels avortaments associats a aquest paràsit, d'utilitat per al control d'aquesta important malaltia.Investigadores de la UAB y de la Universidad de Lleida han observado que infección en laboratorio por el parásito N. caninum en vacas gestantes que s encuentran en el segundo trimestre de gestación es capaz de causar abort imitando lo que sucede en las condiciones de granja. El estudio demuestra que un modelo adecuado para estudiar y tratar de entender la patogenia de lo abortos asociados a este parásito, de utilidad para el control de esta importan enfermedad.Researchers from the Department of Animal Health and Anatomy at the UAB an from the Department of Animal Production at the University of Lleida observe that the infection in laboratory of the N. caninum parasite in pregnant cows alrea in their second trimester can cause abortions, imitating that which has been see to happen under farm conditions. The study demonstrates that it is an adequat model for the study and understanding of the pathogeny of abortion associate with this parasite, and useful for the control of this relevant disease