F J Pallarés

University of Murcia, Murcia, Murcia, Spain

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Publications (98)

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    [Show abstract] [Hide abstract] ABSTRACT: Determination of QTLs for cured ham traits in Duroc by means of genome-wide association study (GWAS). G Ramis1, E Hanenberg2, N Duijvesteijn2, JM Herrero-Medrano2, G Usero3, J Corchero4, JJ Quereda1, L Calvo-Adiego5, AI Rodríguez5, FJ Pallarés6, MB Linares7, MD Garrido7 and A Muñoz1, 3 1Dep. Producción Animal. Universidad de Murcia. Spain, 2 TOPIGS Research center IPG B.V., The Netherlands, 3TOPIGS Ibérica, Spain, 4TOPIGS International., The Netherlands,5Dep. I+D+I Incarlopsa, Spain, 6Dep. Anatomía y Anatomía Patológica Comparadas. Universidad de Murcia, Spain, 7Dep Tecnología de los Alimentos. Universidad de Murcia, Spain. guiramis@um.es Introduction The production of cured ham in Spain represents a very important economic issue in the frame of pork production. The main focus of genetic selection up to the date was related to numeric production. Today, in a “genetic on demand” program the meat quality has increased its importance for costumers. Today the genetic improvement requires of new strategies, including markers assisted selection (MAS). Here we described several Quantitative Trait Loci (QTLs) for cured ham traits in a Spanish Duroc genetic line. Materials and Methods Information from cured hams was available from 249 animals (all TOPIGS Duroc gilts mated with TOPIGS Duroc boars). The parameters studies were: Fat brightness (FTB), Intramuscular fat (IMF), Lean brightness (LNB), Uniformity of color (UNC) and Color intensity (INC), Cured smell intensity (CSI), taste to raw meaty (RWM), Salty taste (SAL), Cured taste (CUR), Hardness (HDN), Chewiness (CWN), Mellowness (MLN), Juiciness (JUN), Total acceptance (ACP), Slice texture average (SLA) and Slice texture standard deviation (SLD). The phenotypic information related to cured ham sensorial meat quality traits was obtained from the Dep. of Food Technology from the University of Murcia (Spain) by means of a sensorial analysis panel, including 2 male and 6 female and it was done in accordance with the ISO 4121 (2003). On the other hand, DNA from 244 animals was isolated and every animal was genotyped using Illumina 60K SNPs chip at GeneSeek (USA). The relation between SNPs markers and productive traits was establishing using a Bayesian variable selection model. It was only consider those regions containing at least two significant SNPs (significance level of Bayes Factor>10) within 1Mb. Results One-hundred and four regions containing at least two significant SNPs (BF>10) within 1Mb were identified for 14 traits. Only 7 QTL regions showed large effects explaining more than 2% of the genetic variation, for ACP, IMF, MLN and HDN. The chromosome, position and number of significant (BF>10) SNPs in the region are shown in table 1. On chromosome (SSC)3, a region showing large effects on several traits together was identified. Five regions showed a significant effect on more than one trait (Table 2). The highest number of effects were found for the trait SAL (29 QTL with BF>10) Table 1. QTLs with a large effect (>2% of σg²), position on chromosome (SSC), start and end (in Mb), and number of significant SNPs (BF>10). QTL SSC Start End Gen. Var explained (%) No. SNPs trait 1 3 13 15 5.42 9 ACP 2 3 16 17 2.88 5 MLN 3 3 17 18 3.23 4 HDN 4 3 17 18 2.23 5 ACP 5 12 46 48 2.50 23 IMF 6 13 189 190 5.57 16 IMF 7 16 63 67 4.33 10 ACP Conclusions and Discussion On the tip of SSC3, Soma et al. (2010) described an effect on meat color in Duroc measured as Minolta values. On SSC12, it has been identified an effect on IMF and marbling score covering the same interval in a Chinese Large-White X Minzhu intercross1. On SSC13, an effect in purebred Spanish Duroc on meat colour Minolta A (redness) of the ham muscle glutaeus medius have been detected2. Moreover, Soma et al. (2010)3 describe an effect in a Japanese Duroc population on IMF in this region (130-195Mb). None of the effect found for SLA trait showed a meaningful overlap with large effects found for the underlying traits. This emphasizes the need to work with detailed phenotypes if the goal is to pinpoint specific region in the genome even with large effects and the underlying functional mutations Acknowledgments The results described in this communication was funded by CDTI “PROCADECO” (nº IDI 20090377); developed by Incarlopsa, Grupo TOPIGS Ibérica, the University of Murcia and TOPIGS Research Center IPG. References 1. Luo et al. 2012. Int J Biol Sci. 8(4):580-95. 2. Gallardo et al. 2012. Anim Gen 43, 800–804 3. Soma et al. 2011. J Anim Sci 89:601-608
    Full-text Conference Paper · Jun 2015
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    F. J. Pallarés · C. Lasa · M. Roozen · G. Ramis
    [Show abstract] [Hide abstract] ABSTRACT: Objectives The purpose of this study was to investigate the efficacy of tylvalosin (Aivlosin Water Soluble Granules, ECO Animal Health) in drinking water for control of Mycoplasma hyopneumoniae (M hyo) on a farm with chronic enzootic pneumonia (EP) problems and high prevalence of mycoplasma-like lesions at slaughter. Design On a 4000-sow farm in the southeast of Spain, 1500 animals of same age were randomly divided into two groups: 900 pigs in the treated group (TG) and 600 pigs in the non-treated control group (CG). TG was medicated for seven days with tylvalosin in drinking water (2.5 mg tylvalosin/kg bodyweight (BW)) at weaning (from 21st to 28th day of life) and a second treatment when moved to finisher barn (from 63rd to 70th day of life). Results In the TG, there was a significant reduction in the severity (P<0.001) and number of animals with lung lesions (P<0.001) compared with CG. TG had an increased average daily gain and decreased average number of days in finishing. TG had a lower average carcase weight, but improved homogeneity. M hyo was not detected by q-PCR in samples, taken from lungs with characteristic EP lesions in the TG (0/9), in contrast to the CG (8/9 positive). Conclusions A strategic medication with Aivlosin at 2.5 mg tylvalosin/kg BW in drinking water for seven days at weaning and when moved to finisher barn significantly reduces mycoplasma-like lung lesions and improves productivity parameters.
    Full-text Article · Jun 2015
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    G Ramis · E Marco · V Magaña · [...] · F J Pallarés
    [Show abstract] [Hide abstract] ABSTRACT: Genetic susceptibility or resistance to diseases is currently drawing increasing attention. This work describes two different breeding herds showing signs of periweaning failure-to-thrive syndrome (PFTS), an emergent swine disease. The disease was diagnosed based on clinical picture and confirmed by histopathology. The possibility of main infectious pathogens was ruled out by immunohistochemistry and PCR. In a simple approach, sires of the affected piglets have been determined using microsatellite paternity analysis, including a healthy group in each case. In each of the two farms, a single boar was found to have sired 45-50 per cent sick animals. Removal of this sire from two farms resulted in a significant decrease in the prevalence of the disease among the offspring, in accordance with other two cases diagnosed, although without including a control group. Since the analysed animals belonged to three different genetic lines, these findings point to the existence of individual genetic susceptibility to this syndrome. British Veterinary Association.
    Full-text Article · Mar 2015
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    Obdulio García-Nicolás · Rubén S Rosales · Francisco J Pallarés · [...] · Francisco J Salguero
    [Show abstract] [Hide abstract] ABSTRACT: Porcine reproductive and respiratory syndrome virus (PRRSV) induces a weak immune response enabling it to persist in different organs of infected pigs. This has been attributed to the ability of PRRSV to influence the induction of cytokine responses. In this study, we investigated the cytokine transcriptional profiles in different compartments of the mediastinal lymph node of pigs infected with three genotype 1 PRRSV strains of differing pathogenicity: the low virulence prototype Lelystad virus (LV), and UK field strain 215–06 and the highly virulent subtype 3 SU1-Bel isolate from Belarus. We have used a combination of laser capture micro-dissection (LCM) followed by real time quantitative PCR (RT-qPCR) and immunohistochemical (IHC) detection of immune cell markers (CD3, CD79a and MAC387) and RT-qPCR quantification of PRRSV and cytokine transcripts. Compared to mock infected pigs, we found a significant downregulation of TNF-α and IFN-α in follicular and interfollicular areas of the mediastinal lymph node from 3 days post-infection (dpi) in animals infected with all three strains. This was accompanied by a transient B cell depletion and T cell and macrophage infiltration in the follicles together with T cell depletion in the interfollicular areas. A delayed upregulation of IFN-γ and IL-23p19 was observed mainly in the follicles. The PRRSV load was higher in all areas and time-points studied in the animals infected with the SU1-Bel strain. This paper describes the first application of LCM to study the cytokine transcript profiles and virus distribution in different compartments of the lymph node of pigs.
    Full-text Article · Mar 2015 · Veterinary Research
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    [Show abstract] [Hide abstract] ABSTRACT: This work was undertaken to evaluate whether a real-time quantitative polymerase chain reaction (qPCR) is as an adequate method for detection and quantification of human-specific DNA elements (Alu gene) in tissues and blood samples of pigs in which human stem cells were engrafted. Real-time qPCR quantification was performed with the use of previously described primers. The human DNA was mixed with different quantities of porcine DNA. The primer concentration and specificity, the qPCR efficiency, the quantification variations due to different porcine DNA concentrations, and the dissociation curve produced by the assay were evaluated. The qPCR proved to be specific, robust, with a reproducible and specific bimodal melting curve. High porcine DNA concentration produced subquantification, especially with low human DNA quantity. However, the assay proved to be useful for the detection of chimeric piglets produced by human cells injected in utero, because the effect caused by the porcine DNA interference was corrected in quantification of human DNA from piglets. Copyright © 2015 Elsevier Inc. All rights reserved.
    Full-text Article · Feb 2015 · Transplantation Proceedings
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    S. B. Morgan · J. P. Frossard · F. J. Pallares · [...] · F. J. Salguero
    [Show abstract] [Hide abstract] ABSTRACT: Porcine reproductive and respiratory syndrome (PRRS) continues to be the most economically important disease of swine worldwide. The appearance of highly pathogenic PRRS virus (PRRSV) strains in Europe and Asia has raised concerns about this disease and initiated increased efforts to understand the pathogenesis. In this study, we have compared the pathology and the virus distribution in tissues of pigs experimentally inoculated with three different genotype 1 PRRSV isolates. Sixty 5-week-old pigs were inoculated intranasally with a) the Lelystad virus (LV), b) a field strain from the UK causing respiratory clinical signs (UK) or c) a highly pathogenic strain from Belarus (BE). Sixteen animals were mock-infected and used as controls. The animals were euthanized at 3, 7 and 35 days post-infection (dpi), and lung and lymphoid tissues collected for histopathological examination and PRRSV detection by immunohistochemistry (IHC). Histopathological lesions consisted of interstitial pneumonia with mononuclear cell infiltrates in the lungs, lymphoid depletion, apoptosis and follicular hyperplasia in the spleen, lymph nodes and tonsil and lymphoid depletion in the thymus. Porcine reproductive and respiratory syndrome virus was detected mainly in monocytes–macrophages. BE-infected animals showed the highest pathological scores and the highest presence of virus at 3 and 7 dpi, followed by the UK field strain and then LV. Moderate lesions were observed at 35 dpi with lesser detection of PRRSV by IHC in each infected group. The highly pathogenic BE strain induced more severe pathology in both lungs and lymphoid organs of pigs compared with the classic field isolate and the prototype LV. The increased severity of pathology was in correlation with the presence of a higher number of PRRSV-infected cells in the tissues.
    Full-text Article · Nov 2014 · Transboundary and Emerging Diseases
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    [Show abstract] [Hide abstract] ABSTRACT: The objective of this study was to determine the effect of crude glycerin (Gly) added to nursery pig diet on nutrient digestibility, the digestive and metabolic status, intestinal morphology and intestinal cytokine expression. A total of 18 male piglets (weaned at 23 days) were used. There were three dietary treatments that differed in the inclusion level of Gly (0, 9 and 18%). On day 14 of the experiment, the animals were weighed and plasma samples were collected before slaughtering. In addition urine, digesta content and intestinal tissues were sampled post mortem. No differences were observed among the tested diets as regards the coefficients of apparent ileal digestibility of DM and CP. The concentration of lactic acid decreased linearly (P<0.05) in the jejunum and ileum segments as the level of Gly increased, although the concentrations of volatile fatty acids in the cecum and colon were not affected. The plasma concentrations of glucose, fructosamine and IGF-1 were not affected. However, urinary glycerol concentrations increased (P<0.01) with increasing levels of Gly. In general, there were no differences in villus height, crypt depth, villus/crypt ratio or number of lymphocytes in the intestinal segments between the different treatments. Nevertheless, the control treatment produced a higher level of goblet cells in the ileum than either of the Gly treatments (P<0.01), while in the jejunum, the number of IgA-secreting cells in the 9% Gly group was higher (P<0.01) than in the control group. There were no differences among the experimental treatments concerning the gene expression of IL-10, IL-12 p40 and TNF-α. Gene expressions of TGF-β, IL-12 p35, IFN-γ and IFN-α were remained unaffected or increased, depending on the intestinal segment and level of Gly addition. In conclusion, the inclusion of Gly at 9 and 18% to the nursery pig diet did not affect nutrient digestibility or plasma metabolites but increased the levels of urinary glycerol, suggesting that metabolic pathways of glycerol utilization became saturated when high levels of Gly are used. In addition, the intestinal cell structure and intestinal cytokine expression might be affected when Gly is included in the feed.
    Full-text Article · Sep 2014 · Livestock Science
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    Obdulio García-Nicolás · Juan José Quereda · Jaime Gómez-Laguna · [...] · Francisco José Pallarés
    [Show abstract] [Hide abstract] ABSTRACT: Porcine Reproductive and Respiratory Syndrome (PRRS) is one of the most economically important diseases of swine. PRRS virus (PRRSV) infection in the pig is characterized by a weak or absent host innate immune response. The underlying mechanisms of PRRSV pathogenesis are still unclear. The analysis of transcript levels represents an alternative to immunoassays for the detection of cytokines that sometimes are difficult to detect due to their low amounts. This study sets out to determine the differences in pathogenesis and the immune response between lung, tonsil, tracheobronchial lymph node (Tb-LN) and retropharyngeal LN (Rf-LN) of PRRSV 2982 strain infected pigs. PRRSV strain 2982 avoided the onset of an effective innate immune response, especially in PRRSV main target (lung) and reservoir (tonsil) organs. PRRSV lead to an impaired expression of IFN-α and TNF-α gene expression, which finally induced a weak and delayed adaptive immune response trough an inefficient IL-12 and IFN-γ expression. Finally, PRRSV replication favoured the expression of the anti-inflammatory IL-10 cytokine in infected pigs.
    Full-text Article · Jul 2014 · Veterinary Immunology and Immunopathology
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    [Show abstract] [Hide abstract] ABSTRACT: Design of a genetic improvement program “on demand”: covering the needs of every costumer A Muñoz1, 2, G Ramis1, E Hanenberg2, JM Herrero-Medrano2, G Usero2, J Corchero4, JJ Quereda1, L Calvo-Adiego5, AI Rodríguez5, FJ Pallarés6, MB Linares7 and MD Garrido7 1Dep. Producción Animal. Universidad de Murcia. Spain, 2 TOPIGS Ibérica, Spain, 3 TOPIGS Research center IPG B.V., The Netherlands, 4TOPIGS International., The Netherlands,5Dep. I+D+I Incarlopsa, Spain, 6Dep. Anatomía y Anatomía Patológica Comparadas. Universidad de Murcia, Spain, 7Dep Tecnología de los Alimentos. Universidad de Murcia, Spain. guiramis@um.es Introduction The genetic improvement has been deeply focused on reproductive and productive performances over the last 50 years. However, the consumers have increased their level of demand regarding meat quality. So, more and more is necessary to develop genetic improvement programs “on demand” of the producer, concerned to cover the consumers requirements. TOPIGS has developed a program on demand focused on meat quality together with zootechnical performances in a Duroc line. Materials and Methods The program was based on 1) accurate data collection of premortem parameters regarding finishing performances, and meat performances such as intramuscular fat by ALOKA, backfat thickness and loin deep. Postmortem lean content by autoFOM was assessed and a wide range of analytical data were recorded regarding fat content, texture, marbling, color, etc. On cured ham samples were analyzed traits such as color, texture, odor, juiciness, etc 2) Genetic analysis to estimate the heritability and the phenotypic and genetic correlations by quantitative approaches, 3) Meat quality analysis, including fresh meat (loin and ham) and cured ham 4) Sensorial analysis of the cured ham to estimate genetic parameters and 4) genome-wide association study (GWAS) using 60K SNPs array to calculate genetic parameters and to establish relation between fresh and cured meat quality and genetic. The meat quality analyses were performed in the Dep. of Food Technology at the University of Murcia. The Illumina’s 60K SNPs chips were analyzed at GeneSeek (USA) after DNA isolation from each animal. The relation between SNPs markers and productive traits was establishing using a Bayesian variable selection model. Calculations for genetic parameters estimation were done using a classical approach by means of the software package ASREML. Results Finally, productive data from 1,590 slaughtered animals included in 30 different slaughtered batches were collected. Carcass classification was obtained from 1,292, meat quality analyses were performed on 249 samples and 60K SNPs data were obtained for 244 animals. As regards heritability estimation, the use of information from 60K SNPs for this purpose showed better results, in terms of standard deviation decrease, than the classical quantitative approach, even using a small population size. The sensorial study and the analytical data showed a good correlation for most of traits related to appearance, taste, texture and acceptance. The GWAS allow identifying eight QTL regions (explaining >2% of σg2) for traits such as Marbling, Ultrasound back fat, Ultrasound Intramuscular fat, Intramuscular fat, and daily gain during test. Talking about cured, seven QTL regions for Total acceptance, Intramuscular fat, Mellowness and Hardness for cured ham traits were identified. Conclusions and Discussion The use of information in vivo related to Intramuscular fat obtained by ALOKA would allow to predict the real intramuscular fat content in carcasses and the heritability of the trait will allow to implement this technic in improvement programs. On the opposite the information of autoFOM post mortem did not supply additional useful information. The GWAS has given a lot of information related to cured ham traits that will help in marked assisted selection (MAS) for lines focused on ham production. The correlation among sensorial traits and analytic data has been medium-high. This information will be used to improve the product on the basis of consumer’s acceptance. The use of sensorial panels, together with molecular analysis is a key point in the design of genetic improvement programs on demand. Acknowledgments The results described in this communication was funded by CDTI “PROCADECO” (nº IDI 20090377); developed by Incarlopsa, Grupo TOPIGS Ibérica, the University of Murcia and TOPIGS Research Center IPG. References 1. Procadeco 2013. Avances, septiembre 2013: 70-74.
    Full-text Conference Paper · Jun 2014
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    G Ramis · E Hanenberg · S Wijga · [...] · A Muñoz
    [Show abstract] [Hide abstract] ABSTRACT: Determination of QTLs for fresh traits in Duroc by means of genome-wide association study (GWAS). G Ramis1, E Henenberg2, S. Wijga2, JM Herrero-Medrano2, G Usero3, J Corchero4, JJ Quereda1, L Calvo-Adiego5, AI Rodríguez5, FJ Pallarés6, MB Linares7, MD Garrido7 and A Muñoz1,3 1Dep. Producción Animal. Universidad de Murcia. Spain,2 TOPIGS Research center IPG B.V., The Netherlands, 3TOPIGS Ibérica, Spain, 4TOPIGS International., The Netherlands,5Dep. I+D+I Incarlopsa, Spain, 6Dep. Anatomía y Anatomía Patológica Comparadas. Universidad de Murcia, Spain, 7Dep Tecnología de los Alimentos. Universidad de Murcia, Spain. guiramis@um.es Introduction The production of cured ham in Spain represents a very important economic issue in the frame of pork production. The main focus of genetic selection up to the date was related to numeric production. Today, in a “genetic on demand” program the meat quality has increased its importance for costumers. Today the genetic improvement requires of new strategies, including markers assisted selection (MAS). Here we described several Quantitative Trait Loci (QTLs) for fresh meat traits in a Spanish Duroc genetic line. Materials and Methods Information from cured hams was available from 248 animals (all TOPIGS Duroc gilts mated with TOPIGS Duroc boars). The parameters studies were: Test daily gain during test (TGR), Ultrasound back fat (BFE), Ultrasound loin depth (LDE), Ultrasound IMF (IMF_LMS), Loin weight (WT_LOIN), weight ham (WT_HAM), Intramuscular fat (IMF), Marbling (MARBLN), Conductivity (CONDUCT), Japanese color (JSCD1), Minolta L value (MINL) Minolta A value (MINA) and Minolta B value (MINB). The phenotypic measurements for TGR, BFE, LDE, IMF_LMS were obtained in the farm, assessed the last three using an ALOKA ultrasound system. On the other hand, to perform the GWAS analysis, DNA from 244 animals was isolated and every animal was genotyped using Illumina 60K SNPs chip at GeneSeek (USA). The relation among SNPs markers and productive traits was establishing using a Bayesian variable selection model. There were only consider those regions containing at least two significant SNPs (significance level of Bayes Factor>10) within 1Mb. Results One hundred and one regions containing at least two significant SNPs (BF>10) within 1Mb were identified for 11 traits in the chromosomes 6, 7, 12, 13, 14 and 15. Among them, eight regions, explaining more than 2% of genetic variation, showed significant relation with different traits. Region 1 (SSC6~66) and region 2 (SSC6~142) were significant for MARBLN and BFE, respectively. For region 3 (SSC7~120), 4 (SSC12~10) and 5 (SSC13~189) evidence was found for genes IMF or marbling. The region 6 (SSC14~5) was related with TGR, and the region 7 (SSC14~123) and 8 (SSC15~152) with BFE. Table 1. QTLs with a large effect (>2% of σg²), position on chromosome (SSC), start and end (in Mb), and number of significant SNPs (BF>10). QTL SSC Start End Gen. Var explained (%) No. SNPs trait 1 6 65.8 66.1 2.9 4 MARBLN 2 6 134.3 137.0 3.5 4 BFE 3 7 118.7 123.2 4.5 8 IMF_LMS 4 12 10. 10.5 3.9 3 MARBLN 5 13 188.7 190.4 6.6 19 IMF 6 14 4.6 4.9 2.5 3 TGR 7 14 121.3 125.6 3.1 12 BFE 8 15 151.4 154.1 2.5 8 BFE Conclusions and Discussion Regions 1 (SSC6~66) and 2 (SSC6~142) are both described in literature for IMF, including candidate genes1. No genes were described in literature in the region 6 (SSC14~5). Maybe this region for TGR should be considered as “false-positive” (3 SNPs only). Region 2 was only significant for BFE, but in literature this region is also linked to IMF2. The correlation of region 7 (SSC14~123) and 8 (SSC15~152) are both described in literature for backfat2 in Duroc. The regions 3, 4 and 5 had been previously related with genes influencing backfat, but not for IMF or MARBLN. Some of these regions have also showed correlation with cured ham traits. In example, region 5 (SSC13~189Mb) was also found for Sensorial IMF and region 8 (SSC14~123Mb) for brightness of fat and loin (LNB, FTB). Acknowledgments The results described in this communication was funded by CDTI “PROCADECO” (nº IDI 20090377); developed by Incarlopsa, Grupo TOPIGS Ibérica, the University of Murcia and TOPIGS Research Center IPG. References 1. Árnyasi et al. 2006. J Anim Breed Genet. 123:198–203 2. Gallardo et al. 2012. Anim Gen 43, 800–804
    Full-text Conference Paper · Jun 2014
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    Full-text Conference Paper · Jun 2014
  • Juan Seva · Jose M. Sanes · Guillermo Ramis · [...] · Francisco J. Pallares
    [Show abstract] [Hide abstract] ABSTRACT: This study reports the performance of the single intradermal tuberculin (SIT) test and the interferon-gamma (IFN-γ) assay for M. bovis in a cattle herd with high prevalence of paratuberculosis (PTB). A total of 58/350 animals were selected for necropsy based on one or more of the following criteria: positive to SIT, IFN-γ, a breeding cow that seroconverted to PTB and showed signs compatible with a wasting disease. Infection status was determined by post mortem diagnostic tests that included histopathology examination and mycobacterial cultures coupled with PCR identification for M. bovis and M. avium subsp. paratuberculosis (MAP). In 7/58 animals primary tuberculosis (TB) lesions, affecting only the retropharyngeal and/or mediastinal lymph nodes, were found; 3/7 animals were found SIT positive. PTB was confirmed in 35/58 animals, of which 30 had seroconverted and 14 had typical clinical signs. 45/58 animals were IFN-γ+ using the most stringent criterion (cut-off point≥0.05); however, IFN-γ test was only positive in 33 animals when using a higher threshold (cut-off point≥0.1). Three animals co-infected also showed extensive TB and diffuse PTB lesions. These results show that the combined use of SIT and IFN-γ, as interpreted using official guidelines, detected all confirmed cases of TB. Individually, the sensitivity of the SIT was inadequate to diagnose TB-positive animals with an advanced stage of PTB. The large number of IFN-γ+ animals with no visible TB lesion could be due, in part, to some protection conferred by prior infection with MAP
    Article · Jun 2014 · Veterinary Microbiology
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    Full-text Article · Jan 2014 · Journal of Comparative Pathology
  • RS Rosales · FJ Pallarés · J Gough · [...] · F.J. Salguero
    Article · Jan 2014 · Journal of Comparative Pathology
  • S.B. Morgan · F.J. Pallares · J. Gough · [...] · F.J. Salguero
    Article · Jan 2014 · Journal of Comparative Pathology
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    [Show abstract] [Hide abstract] ABSTRACT: The polarization into M1 and M2 macrophages (MΦ) is essential to understand MΦ function. Consequently, the aim of this study was to determine the impact of IFN-γ (M1), IL-4 (M2) and IFN-β activation of MΦ on the susceptibility to genotype 1 and 2 porcine reproductive respiratory syndrome (PRRS) virus (PRRSV) strains varying in virulence. To this end, monocyte-derived MΦ were generated by culture during 72h and polarization was induced for another 24h by addition of IFN-γ, IL-4 or IFN-β. MΦ were infected with a collection of PRRSV isolates belonging to genotype 1 and genotype 2. Undifferentiated and M2 MΦ were highly susceptible to all PRRSV isolates. In contrast, M1 and IFN-β activated MΦ were resistant to low pathogenic genotype 1 PRRSV but not or only partially to genotype 2 PRRSV strains. Interestingly, highly virulent PRRSV isolates of both genotypes showed particularly high levels of infection compared with the prototype viruses in both M1 and IFN-β-treated MΦ (p<0.05). This was seen at the level of nucleocapsid expression, viral titres and virus-induced cell death. In conclusion, by using IFN-γ and IFN-β stimulated MΦ it is possible to discriminate between PRRSV varying in genotype and virulence. Genotype 2 PRRSV strains are more efficient at escaping the intrinsic antiviral effects induced by type I and II IFNs. Our in vitro model will help to identify viral genetic elements responsible for virulence, an information not only important to understand PRRS pathogenesis but also for a rational vaccine design. Our results also suggest that monocyte-derived MΦ can be used as a PRRSV infection model instead of alveolar MΦ, avoiding the killing of pigs.
    Full-text Article · Nov 2013 · Virus Research
  • Conference Paper · Sep 2013
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    [Show abstract] [Hide abstract] ABSTRACT: This paper presents an optimized procedure for assessing an immune-mediated cytotoxicity, produced after the addition of human and baboon serum to transgenic porcine fibroblasts. This procedure is performed with the xCELLigence Real-Time Cell Analyzer (RTCA). The xCELLigence system measures the impedance variations in the culture media of a 96-well microelectronic plate, and shows the changes in cell number and morphology in a real-time plot. However, different factors need to be optimized before developing an RTCA assay. Thus, we studied the influence of several variables, such as the number of cells seeded, the time the cells were allowed to grow before the tests, the serum concentration and the addition of rabbit complement. The findings were confirmed by the WST-1 classical cytotoxicity test. The results showed that 7.5 × 10(3) cells seeded per well produced the adequate CI in 10 h. The area under the curve and the CImin versus concentration values showed a very high correlation index (r(2) = 0.966 and r(2) = 0.92 for the first 50 h after challenge, respectively), proving that CI variations are directly proportional to the quantity of serum added. The addition of complement resulted in lower CImin values. Therefore, both the cytolysis level with and without exogenous complement addition had to be assessed. There was a high correlation between the relative cytotoxicity assessed by WST-1 and the CI obtained by RTCA when exogenous complement was not added (r(2) = 0.827; p < 0.001). The correlation was average when rabbit complement was added (r(2) = 0.523; p = 0.046). In conclusion, culture conditions have an important influence on RTCA cytotoxicity assays.
    Full-text Article · Jul 2013 · Biomedical Microdevices
  • [Show abstract] [Hide abstract] ABSTRACT: A 12-year-old, male, fox terrier dog presented with an abnormal gait of the left pelvic limb. Computed tomography revealed a large, homogeneous, hypoattenuating, noncontrast enhancing mass within the left epaxial muscles that invaded the L5-6 vertebral canal and caused spinal cord compression. Imaging findings were consistent with an infiltrative lipoma. The mass was removed and a left hemilaminectomy was performed in the affected area. Histopathology confirmed the mass to be an infiltrative lipoma. The dog recovered and regained neurologic function within 2 weeks. Computed tomography assisted preoperative planning by characterizing the shape, size, and location of the mass.
    Article · Apr 2013 · Veterinary Radiology & Ultrasound
  • Conference Paper · Jan 2013