Technical Note: A new device for cervical insemination of sheep - design and field test.

Deep semen deposition, avoiding retrograde flow, lesions and stress, has proved to be very important in the success of sheep AI. The objective of the present study was to develop a new, suitable anti-retrograde flow device for sheep cervical AI (DARIO) that enables deep deposition of semen into the cervix without any modifications to the procedures currently used, and to compare the fertility, fecundity, and prolificacy rates between DARIO and a traditional catheter. Field tests were performed on 16 farms actively participating in the non-profit National Association of Rasa Aragonesa Breeders´ genetic selection scheme and where sheep management was similar. A total of 242 AI lots were considered, including 1,299 ewes; 126 lots (662 ewes) were inseminated using DARIO, and 116 lots (637 ewes) using a traditional commercially-available catheter (control group). Several factors affecting AI results were included in the model for mean comparison between DARIO and control groups (farm and ram as random factors; catheter, year and photoperiod as fixed effects; catheter × photoperiod interaction). The type of catheter had a significant effect on fertility ( < 0.01) and fecundity rates ( < 0.01) but no significant effect was detected on the prolificacy rate ( = 0.45). For fertility rate (percentage of ewes lambing after AI), means ± SE for DARIO and control groups were 59.44 ± 2.13% and 49.60 ± 2.48%, respectively; for fecundity rates, means ± SE for DARIO and control groups were 0.99 ± 0.04 and 0.82 ± 0.05 lambs/inseminated ewe, respectively, and, for prolificacy rates, means ± SE for DARIO and control groups were 1.68 ± 0.04 and 1.63 ± 0.04 lambs/ewe lambing, respectively. Fertility rate was greater in the decreasing photoperiod ( = 0.01). Significant effects were found for both year ( < 0.05) and farm ( < 0.01) on fertility, fecundity, and prolificacy rates. Neither ram nor catheter × photoperiod showed any significant effects on the variables investigated ( > 0.05). Overall, the use of DARIO instead of the traditional commercially-available catheter increased both fertility and fecundity rates; the marginal mean differences were 9.05 pregnant ewes per 100 inseminated and 0.15 lambs per inseminated ewe, respectively.

Acute physiological responses to castration-related pain in piglets: the effect of two local anesthetics with or without meloxicam.

Methods to reduce castration-related pain in piglets are still issues of concern and interest for authorities and producers. Our objectives were to estimate the effectiveness of two protocols of local anesthesia (lidocaine and the combination of lidocaine+bupivacaine) as well as the use of meloxicam as a postoperative analgesic in alleviating castration-related pain, measured by acute physiological responses. Eight groups (15 piglets/group) were included in the study: (1) castration without anesthesia or analgesia, without meloxicam (TRAD WITHOUT), (2) castration without anesthesia or analgesia, but with meloxicam (TRAD WITH), (3) handling without meloxicam (SHAM WITHOUT), (4) handling with meloxicam (SHAM WITH), (5) castration after local anesthesia with lidocaine but without meloxicam (LIDO WITHOUT), (6) castration after local anesthesia with lidocaine and meloxicam (LIDO WITH), (7) castration after local anesthesia with lidocaine+bupivacaine without meloxicam (LIDO+BUPI WITHOUT), (8) castration after local anesthesia with lidocaine+bupivacaine and meloxicam (LIDO+BUPI WITH). Acute physiological responses measured included skin surface temperature and serum glucose and cortisol concentrations. On days 4 and 11 post-castration BW was recorded and average daily gain was calculated over this period. Furthermore, piglet mortality was recorded over the 11-day post-castration period. Administration of local anesthetic or meloxicam did not prevent the decrease in skin surface temperature associated with castration. Lidocaine reduced the increase in glucose concentration associated with castration. For castrated pigs, the joint use of lidocaine and meloxicam caused a significant decrease in cortisol concentration; the combination of intratesticular lidocaine and bupivacaine did not seem to be more effective than lidocaine alone. No effect of treatments on mortality and growth were detected.

Frey syndrome: factors influencing the time to event.

Frey syndrome is a common complication after parotidectomy. The time from surgery to disease onset may be quite long; therefore, a time-to-event analysis was performed for the occurrence of this syndrome post-parotidectomy. Three hundred and thirty-four patients who underwent a parotidectomy between January 2002 and November 2012 were identified (retrospective study). Of these patients, 102 developed Frey syndrome post-surgery and 232 did not. The time-to-onset analysis enabled us to estimate the risk ratio associated with different types of parotid gland tumours, various parotidectomy procedures, and repeat parotidectomy, which is useful for predicting preoperative and surgical risk. The risk of developing Frey syndrome was lower in patients with malignant tumours than in those with benign tumours (risk ratio 0.351, 95% confidence interval (CI) 0.155-0.594). Risk ratios for lumpectomy PA (pre-auricular area), superficial parotidectomy, and total parotidectomy with respect to lumpectomy T (tail) were 4.378 (95% CI 1.168-16.410), 8.040 (95% CI 3.286-19.670), and 8.174 (95% CI 3.076-21.723), respectively. Repeat parotidectomy also increased the risk of developing Frey syndrome (risk ratio 3.214, 95% CI 1.547-6.678). No effect of the use of a superficial muscular aponeurotic system (SMAS) flap on the risk of developing Frey syndrome was detected (P=0.888).

Canine stage 1 periodontal disease: a latent pathology.

To evaluate the potential health issues associated with periodontal disease (PD) in dogs, 1004 teeth from 25 dogs were examined. The dogs were randomly selected, aged 2-14 years, and had at least 95% of their teeth at the first PD stage. Significant positive correlations between plaque grade (PG) and gum inflammation, gingival regression, periodontal pocket, age and serum alanine aminotransferase (ALT) activity were identified. In contrast, PG was negatively correlated to total platelet count. Altogether, these findings suggest that prevention and therapy at the first PD stages can have an important impact on the general health condition of dogs.

Anatomical and genetic study of an ancient animal tooth showing brachyodont and hypsodont mixed taxonomical characteristics.

A non-human dental piece was found in a Roman Empire tomb dated the 3rd century A.C. in Zaragoza (Spain). The morphology of this piece showed mixed brachyodont (carnivores) and hypsodont (herbivores) characteristics. As a result, the taxonomical assignation of the piece was impossible. Therefore, a protocol based on the DNA sequence of the cytochrome c oxidase subunit 1 mitochondrial region (COI) was applied. For this purpose, a pair of primers able to amplify this region in a large variety of animals was designed. The results point to a species of the Genus Bos (Family Bovidae). This assignation was later confirmed by these quencing of a short fragment of the mitochondrial D-loop region. A complete morphological description of the tooth is presented together with the DNA sequence study and comparison protocol.

A presumptive new locus for autosomal dominant hypercholesterolemia mapping to 8q24.22.

Molecular testing of patients with autosomal dominant hypercholesterolemia (ADH) fails to detect a causal functional mutation in 15.25% of subjects. We studied an ADH pedigree in which known ADH-causing genes (LDLR, APOB and PCSK9) were excluded. Genome-wide analysis on 15 family members detected significant association for ADH and dbSNP RS ID rs965814 (G/A), located in 8q24.22 cytoband. ADH was significantly associated to rs965814 G allele (p < 0.05) in a case-control study based on 200 unrelated ADH subjects without LDLR or APOB gene defects and 198 normolipidemic controls. We chose 24 markers for a detailed analysis of 8q24.22 cytoband, now based on an extended set of family members (21 individuals). One particular 24 marker haplotype was significantly associated to both higher total and low-density lipoprotein-cholesterol concentrations. Similar results were found for a shorter haplotype, composed of the distal six markers from the complete haplotype. Therefore, a presumptive new locus for ADH could be located in 8q24.22 cytoband, a region not previously linked or associated to ADH.

Haplotype analyses, mechanism and evolution of common double mutants in the human LDL receptor gene.

Familial hypercholesterolemia (FH), an autosomal dominant inherited disorder resulting in increased levels of circulating plasma low-density lipoprotein (LDL), tendon xanthomas and premature coronary artery disease (CAD), is caused by defects in the LDL receptor gene (LDLR). Three widespread LDLR alterations not causing FH (c.1061-8T>C, c.2177C>T and c.829G>A) and one mutation (c.12G>A) with narrow geographical distribution and thought to cause disease were investigated. In an attempt to improve knowledge on their origin, spread and possible selective effects, estimations of the ages of these variants (t generations) and haplotype analysis were performed by genotyping 86 healthy individuals and 98 FH patients in Spain for five LDLR SNPs: c.81T>C, c.1413G>A, c.1725C>T, c.1959T>C, and c.2232G>A; most patients carried two of these LDLR variants simultaneously. It was found that both the c.1061-8T>C (t = 54) and c.2177C>T alterations (t = 62) arose at about the same time (54 and 62 generations ago, respectively) in the CGCTG haplotype, while the c.12G>A mutation (t = 70) appeared in a CGCCG haplotype carrying an earlier c.829G>A alteration (t = 83). The estimated ages of selectively neutral alterations could explain their distribution by migrations. The origin of the c.12G>A mutation could be in the Iberian Peninsula; despite its estimated age, a low selective pressure could explain its conservation in Spain from where it could have spread to China and Mexico, since the sixteenth century through the Spanish/Portuguese colonial expeditions.

Ovine heritable arthrogryposis multiplex congenita with probable lethal autosomal recessive mutation.

A novel congenital syndrome in lambs is described that is characterized by: (1) kyphoscoliosis (severe curvature, hunching and twisting of the thoracic spine, with associated abnormalities of the ribs and sternum), (2) distal arthrogryposis of the carpal and tarsal joints, and (3) cleft hard and soft palate or palatoschisis (a median fissure of the palate). Male and female lambs are affected equally and in multiple births there may be only one affected lamb. The affected lambs are born full term, but die shortly after birth because they are unable to start breathing. Affected lambs have slightly reduced bodyweight (due to low muscle mass) compared with normal newborn lambs of the same flock. The syndrome is remarkably similar to the recently described bovine heritable arthrogryposis multiplex congenita in the Angus breed. Genetic analysis of the flock by means of DNA microsatellite markers, together with study of the reproductive management of the flock, allowed us to discount an environmental aetiology (e.g. intoxications or infections). In contrast, evidence supports an autosomal recessive inherited disease for which the name ovine heritable arthrogryposis multiplex congenita is proposed.

A case of ovine female XY sex reversal syndrome not related to anomalies in the sex-determining region Y (SRY).

The human XY gonadal sex reversal is a rare disease. Primary amenorrhea, lack of secondary sexual characteristics and the possible existence of streak gonads are the most common signs of this syndrome. Five cases of this syndrome have been described in the last 42 years in bovines. However, to the knowledge of the authors, no similar descriptions are available in sheep. In the present report, we are presenting the case of a 4 year old ewe 2n = 54,XY showing primary sterility and total absence of masculinization signs. The promoter and the complete coding segment of the sex determining region Y (SRY) were sequenced and found to be absolutely normal. The presence of the testis-specific protein gene (TSPY) was also verified by means of PCR. Necropsy findings include short vagina, lack of uterus and complete absence of masculine remains. Streak gonads concordant with previous descriptions in humans were also found.

Susceptibilidad a antimicrobianos ytipificación molecular de Enterococcusfaecium aislados de muestras de origenhumano, animal y ambiental en Gran Canaria / Antimicrobial susceptibility and molecular typing of Enterococcus faecium isolated from humans, chickens and environment in Canary Islands (Spain)

El estudio comparativo de los perfiles de sensibilidadfrente a antimicrobianos y la tipificación molecular de los aisladosde Enterococcus de diferentes orígenes pueden proporcionarinformación importante para el análisis epidemiológicode las infecciones causadas por este género bacteriano.Se estudiaron aislados clínicos y se tomaron muestras de hecesde humanos (personas hospitalizadas y voluntarios sanos),heces de aves y muestras ambientales. Se obtuvieron 68 aisladosde E. faecium, de los cuales 43 procedían de humanos,5 de aves y 20 de aguas. Se estudiaron los patrones y los mecanismosde resistencia a antibióticos y se caracterizaron losaislados mediante sus perfiles al aplicar electroforesis encampo pulsante (PFGE). Se usó la reacción en cadena de la polimerasa(RCP) para detectar la presencia de ocho genes de resistenciaa antibióticos aminoglicósidos. Se observaron diferenciasen los porcentajes de resistencia a antimicrobianos enlas muestras clínicas y no clínicas. Todos los aislados fueronsensibles a vancomicina y teicoplanina. Se detectaron cuatrogenes de resistencia a aminoglicósidos, siendo los más frecuentesant(6)-Ia y aph(3’)-IIIa. La presencia de aislados resistentesa gentamicina en los que no se detectaron genes deresistencia mediante RCP sugiere que pueden existir otros genesde resistencia adicionales. La alta frecuencia de aisladosresistentes a ampicilina entre los enterococos procedentes demuestras clínicas, y el hecho de que varios aislados compartanel mismo perfil de PFGE parece sugerir la presencia de unacepa de E. faecium resistente a ampicilina endémica en nuestrohospital (AU)

Comparative studies on antimicrobial susceptibilitypatterns and molecular typing of Enterococcus isolates ofdifferent origins provides valuable information concerningthe epidemiology of enterococcal infections. We analyzedclinical isolates and we surveyed faecal samples of humans(hospitalised patients and healthy volunteers), faecal samplesof poultry and environmental samples. A total of 68E. faecium isolates were obtained: 43 from humans, 5 frompoultry and 20 from water. We compared the antibiotic resistancepatterns and pulsed field gel electrophoresis (PFGE)profiles of these strains.We used polymerase chain reaction(PCR) to examine them for the presence of 8 aminoglycosideresistance genes. Differences among percentages of antimicrobialresistance between clinical and non clinicalisolates were found. All enterococci were susceptible to vancomycinand teicoplanin. Four aminoglycoside resistancegenes were detected, most frequently ant(6)-Ia andaph(3’)-IIIa. Presence of isolates resistant to gentamicin butnegative for all genes tested suggest that additional resistancegenes may exist. VRE are still rare inside and outsidehospitals in Gran Canaria (Spain). The high frequency ofampicillin resistance among clinical enterococci and the factthat several isolates share the same PFGE type were isolatedfrom different wards of our hospital suggest that ampicillinresistantE. faecium are endemic in our Hospital (AU)

[Antimicrobial susceptibility and molecular typing of Enterococcus faecium isolated from humans, chickens and environment in Canary Islands (Spain)]. / Susceptibilidad a antimicrobianos y tipificación molecular de Enterococcus faecium aislados de muestras de origen humano, animal y ambiental en Gran Canaria.

Comparative studies on antimicrobial susceptibility patterns and molecular typing of Enterococcus isolates of different origins provides valuable information concerning the epidemiology of enterococcal infections. We analyzed clinical isolates and we surveyed faecal samples of humans (hospitalised patients and healthy volunteers), faecal samples of poultry and environmental samples. A total of 68 E. faecium isolates were obtained: 43 from humans, 5 from poultry and 20 from water. We compared the antibiotic resistance patterns and pulsed field gel electrophoresis (PFGE) profiles of these strains.We used polymerase chain reaction (PCR) to examine them for the presence of 8 aminoglycoside resistance genes. Differences among percentages of antimicrobial resistance between clinical and non clinical isolates were found. All enterococci were susceptible to vancomycin and teicoplanin. Four aminoglycoside resistance genes were detected, most frequently ant(6)-Ia and aph(3')-IIIa. Presence of isolates resistant to gentamicin but negative for all genes tested suggest that additional resistance genes may exist. VRE are still rare inside and outside hospitals in Gran Canaria (Spain). The high frequency of ampicillin resistance among clinical enterococci and the fact that several isolates share the same PFGE type were isolated from different wards of our hospital suggest that ampicillin-resistant E. faecium are endemic in our Hospital.

Microsatellite markers for the analysis of genetic variability and relatedness in red-legged partridge (Alectoris rufa) farms in Spain.

The usefulness of several microsatellites in the management of Alectoris rufa restocking farms is evaluated in the present report. We analysed seven microsatellite loci in 147 randomly chosen individuals from four Spanish farms. Global F(IS) values were not significant. Slight genetic differentiation was only found between Farms 1 and 4. The global first and second parent exclusionary powers were 0.742 and 0.930, respectively. Microsatellite analysis can estimate the genetic relatedness between pairs of individuals by means of the Identity index. Since genealogies are unknown in most farms, pairwise Identity index values proved to be helpful in the management of matings, and the improvement of reproductive success and fitness of the descendants. Mean Identity index values showed that individuals within farms were not genetically more related than expected under random mating. Variance of the Identity index values suggest the absence of closed familial groups in these farms.

Scrapie resistance alleles are not associated with lower prolificity in Rasa Aragonesa sheep.

Scrapie is a prion disease characterised by the accumulation of the pathological associated form of cellular prion protein (PrP(SC)) in the central nervous system. Susceptibility to scrapie is associated with polymorphism in the ovine prion protein (PrP) gene. The European Union has implemented scrapie control programs, relying on selective breeding for scrapie resistance; the use of ARR-carrier and the exclusion of VRQ-carrier were recommended. In this study, 4323 individuals from Rasa Aragonesa Sheep breed were genotyped for the PrP gene and the individual estimated breeding values (EBV) for prolificity were calculated. Most represented PrP alleles do not work against prolificity. Only a significant association between VRQ/VRQ genotype and a lower EBV was observed (p = 0.027, eta2 = 0.002). Therefore, avoiding reproduction of VRQ/VRQ individuals would not cause negative effect regarding prolificity.

Phylogenetic relationships among Spanish goats breeds.

We partially sequenced the mitochondrial hypervariable region 1 (HVR1) in 60 goats belonging to six Spanish breeds. The analysis of these and previously published sequences reveals a weak phylogeographical structure in the Iberian Peninsula breeds. Individuals from a single breed did not group into a single cluster. Furthermore, individuals from different breeds often shared single phylogenetic tree branches after UPGMA analysis. This could reflect the non-existence of breed isolation because of traditional seasonal pastoralism and annual long-distance migrations. Three goats belonging to the C maternal lineage were found, demonstrating a wider than previously thought distribution for this lineage.

Perforación duodenal neonatal espontánea / Spontaneous duodenal perforation in the neonate

No disponible

Analysis of microsatellites and paternity testing in Rasa Aragonesa sheep.

Four microsatellite loci (MAF50, MAF18, OarFCB20 and MCM527) were studied in Rasa Aragonesa sheep in order to evaluate their use in paternity testing. Several population characteristics were estimated [allele frequencies. effective allele number (Ne), polymorphism informative content (PIC) and probability of excluding wrong paternities (Pe)]. In 32 randomly chosen individuals, four alleles were detected for MAF50, with 2.55 effective alleles, 0.58 PIC and 0.35 Pe. For MAF18, five alleles were identified, with 2.99 effective alleles, 0.51 PIC and 0.32 Pe. For oarFCB20, 10 alleles were observed, with 6.06 effective alleles, 0.82 PIC and 0.68 Pe. Finally, for MCM527, six alleles were found, with 3.75 effective alleles, 0.69 PIC and 0.50 Pe. When these loci were used together with serum transferrin locus, Pe rose to 97.20 per cent. Field trials confirmed the real usefulness of these techniques.

Climatic effects on Salmonella survival in plant and soil irrigated with artificially inoculated wastewater: preliminary results.

The use of wastewater to replace other water resources for irrigation is highly dependent on whether the health risk and environmental impacts entailed are acceptable or not. Total count and species of microorganisms found in wastewater vary widely because of climatic conditions, season, population sanitary habits and disease incidence. Salmonella, one of the genera associated with waterborne diseases, lives in the intestine. Thus, it is widely accepted that they have a limited survival period under environmental conditions. Wastewater management practices and the ability of Salmonella to survival under field conditions would determine the health risk associated with its presence in wastewater. Although chlorination is widely used, there are situations in which Salmonella is able to survive the sudden stress imposed by this technique. The aim of this experiment was to contribute to the study of the climatic and soil effects on pathogen survival under agricultural field conditions in order to assess which were the best wastewater management practices from both health and economic points of view. Five pots filled with soil seeded with Medicago sativa and an automatic weather station were used. A secondary effluent was artificially inoculated with Salmonella. In addition, open plates (filled with sterilised soil) and ultraviolet radiation isolated plates (filled with non-sterilised soil) were used. As soil heat emission contributes to the environmental conditions around the bacteria, standardised meteorological temperature data had to be carefully used in the bacterial survival studies under agricultural conditions. Radiation was the main cause of Salmonella mortality as its effect was more important than natural soil bacteria competence. Higher reduction of Salmonella counts could have been associated with longer spring days. Soil was able to effectively remove Salmonella. Subsurface drip irrigation methods could provide an effective tool to prevent health risk associated with wastewater irrigation.