N. Carolino

Genetic diversity in native and commercial breeds of pigs in Portugal assessed by microsatellites

Population structure and genetic diversity in the Portuguese native breeds of pigs Alentejano (AL), Bísaro (BI), and Malhado de Alcobaça (MA) and the exotic breeds Duroc (DU), Landrace (LR), Large White (LW), and Pietrain were analyzed by typing 22 microsatellite markers in 249 individuals. In general, the markers used were greatly polymorphic, with mean total and effective number of alleles per locus of 10.68 and 4.33, respectively, and an expected heterozygosity of 0.667 across loci. The effective number of alleles per locus and expected heterozygosity were greatest in BI, LR, and AL, and least in DU. Private alleles were found in 9 of the 22 markers analyzed, mostly in AL, but also in the other breeds, with the exception of LW. The proportion of loci not in Hardy-Weinberg equilibrium in each breed analyzed ranged between 0.23 (AL) and 0.41 (BI, LW, and Pietrain), mostly because of a less than expected number of heterozygotes in those loci. With the exception of MA, all breeds showed a significant deficit in heterozygosity (F(IS); P < 0.05), which was more pronounced in BI (F(IS) = 0.175) and AL (F(IS) = 0.139), suggesting that inbreeding is a major concern, especially in these breeds that have gone through a genetic bottleneck in the recent past. The analysis of relationships among breeds, assessed by different methods, indicates that DU and AL are the more distanced breeds relative to the others, with the closest relationship being observed between LR and MA. The degree of differentiation between subpopulations (F(ST)) indicates that 0.184 of the total genetic variability can be attributed to differences among breeds. The analysis of individual distances based on allele sharing indicates that animals of the same breed generally cluster together, but subdivision is observed in the BI and LR breeds. Furthermore, the analysis of population structure indicates there is very little admixture among breeds, with each one being identified with a single ancestral population. The results of this study confirm that native breeds of pigs represent a very interesting reservoir of allelic diversity, even though the current levels of inbreeding raise concerns. Therefore, appropriate conservation efforts should be undertaken, such as adopting strategies aimed at minimizing inbreeding, to avoid further losses of genetic diversity.

Effect of hot-iron disbudding on behaviour and plasma cortisol of calves sedated with xylazine

We investigated cortisol and behaviour for the first hour after hot-iron disbudding of calves aged 37+/-4 days: disbudded after i/m xylazine (n=10); disbudded after i/m xylazine and regional anaesthesia with lidocaine (n=10); sham-disbudded after xylazine and lidocaine (n=11); sham-disbudded after i/m saline and lidocaine (n=10). Xylazine-treated groups had higher cortisol than saline-treated animals and showed no differences among them at any time. Sham-disbudded calves with xylazine had lower cortisol at 60 min compared with all other times. Xylazine-alone disbudded calves struggled more during the procedure than all other groups. Xylazine-alone disbudded calves showed more ear-flicks at 10, 25 and 40 min and head-shakes at 40 min than all other groups. We conclude that cortisol should not be used as an indicator of pain in disbudded calves while under the sedative effect of xylazine and that some behaviours during and after the procedure are useful in showing that xylazine alone does not control hot-iron disbudding pain.

Inbreeding depression on beef cattle traits: Estimates, linearity of effects and heterogeneity among sire-families

Records from up to 19054 registered cows and 10297 calves in 155 herds of the Alentejana cattle breed were used to study the effects of individual (Fi) and maternal (Fm) inbreeding on reproductive, growth and carcass traits, as well as assessing the importance of non-linear associations between inbreeding and performance, and evaluating the differences among sire-families in the effect of Fi and Fm on calf weight at 7 months of age (W7M). Overall, regression coefficients of performance traits on inbreeding were small, indicating a minor but still detrimental effect of both Fi and Fm on most traits. The traits with the highest percentage impact of Fi were total number of calvings through life and calf weight at 3 months of age (W3M), followed by longevity and number of calves produced up to 7 years, while the highest effect of Fm was on W3M. Inbreeding depression on feed efficiency and carcass traits was extremely small and not significant. No evidence was found of a non-linear association between inbreeding and performance for the traits analyzed. Large differences were detected among sire-families in inbreeding depression on W7M, for both Fi and Fm, encouraging the possibility of incorporating sire effects on inbreeding depression into selection decisions.

Serological evidence of exposure to Coxiella burnetii in sheep and goats in central Portugal.

The recent outbreak of Q fever in The Netherlands warned European health authorities of the need of studying Coxiella burnetii. In Portugal, little is known about C. burnetii infection in animals. A cross-sectional study was designed to investigate the exposure to C. burnetii in sheep and goats in the Central region of Portugal, estimating the herd and individual prevalence. A serosurvey was conducted in a two levels random sampling of 89 herds and 460 animals. Individual blood samples were collected from animals older than 6 months, and specific antibodies anti-C. burnetii were detected by ELISA testing. Results showed a global herd prevalence of 32.6% (95% CI: 23.1-42.1%). Herd prevalence was higher in mixed herds (38.5%; 95% CI: 12-65%) and in sheep herds (37.5%; 95% CI: 21-54%) than in goat herds (28.8%; 95% CI: 17-41%). Global individual prevalence was estimated at 9.6% (95% CI: 6.9-12.2%), and it was higher in goats (10.4%; 95% CI: 7.8-13%) than in sheep (8.6%; 95% CI: 5.8-11.4%). Sample positive percentages (S/P) ranged from 41.5% to 185.9%. S/P percent higher than 100 was found in 18.2% (8/44) of sera from distinct herds. Positive results were significantly associated with goats, older animals and larger herds. These results revealed the presence of C. burnetii in small ruminants evidencing their potential role in the infection cycle.

Implementation of a parentage control system in Portuguese beef-cattle with a panel of microsatellite markers

A study was conducted to assess the feasibility of applying a panel of 10 microsatellite markers in parentage control of beef cattle in Portugal. In the first stage, DNA samples were collected from 475 randomly selected animals of the Charolais, Limousin and Preta breeds. Across breeds and genetic markers, means for average number of alleles, effective number of alleles, expected heterozygosity and polymorphic information content, were 8.20, 4.43, 0.733 and 0.70, respectively. Enlightenment from the various markers differed among breeds, but the set of 10 markers resulted in a combined probability above 0.9995 in the ability to exclude a random putative parent. The marker-set thus developed was later used for parentage control in a group of 140 calves from several breeds, where there was the suspicion of possible faulty parentage recording. Overall, 76.4% of the calves in this group were compatible with the recorded parents, with most incompatibilities due to misidentification of the dam. Efforts must be made to improve the quality of pedigree information, with particular emphasis on information recorded at the calfs birth.

Effect of a quadrivalent vaccine against respiratory virus on the incidence of respiratory disease in weaned beef calves

Abstract We investigated the effect of vaccination of male beef calves (mean age±S.D.: 158±31days) against bovine herpes virus (BHV-1 or IBR virus), bovine respiratory syncitial virus (BRSV), bovine viral diarrhea (BVD) virus and para-influenza (PI3) virus on the incidence of respiratory disease during the first forty days after weaning and entering a feed-lot in Portugal. In May 2003, Mertolenga, Preta and mixed-breed calves from 10 different beef herds, were systematically assigned (by order of entrance in a chute) to two treatment groups, before moving to a common feed-lot. One hundred and twenty five male calves were vaccinated with a quadrivalent vaccine (Rispoval 4®) and revaccinated after 21–27 days while 148 herdmates were injected with saline (0.9% NaCl) on the same occasions. The incidence and severity of clinical cases of “bovine respiratory disease” (BRD) were evaluated every day during the first 40 days after entering the feed-lot. Morbidity (3% vs. 14%) and mortality (0% vs. 4%) due to BRD were significantly lower in the vaccinated group. Ten days after revaccination, the calves were treated with an antimicrobial – ending the study – after an outbreak of BRD caused a high incidence of disease in the non-vaccinated group. In conclusion, our results showed that Rispoval 4®, a quadrivalent vaccine against respiratory viruses, under field conditions, reduces morbidity and mortality due to BRD in beef calves after weaning.

Fat area and lipid droplet morphology of porcine oocytes during in vitro maturation with trans -10, cis -12 conjugated linoleic acid and forskolin

Lipid droplets (LD) in porcine oocytes form a dark mass reaching almost all cytoplasm. Herein we investigated changes in fat areas, cytoplasmic tone and LD morphology during in vitro maturation (IVM) of porcine oocytes cultured with 100 μM trans-10, cis-12 conjugated linoleic acid (t10,c12 CLA) or 10 μM forskolin at different time periods. Four groups were constituted: control, excipient, t10,c12 CLA and forskolin, with drugs being supplemented during 44 to 48 h and the initial 22 to 24 h in Experiments 1 and 2, respectively. In Experiment 3, forskolin was supplemented for the first 2 h. Matured oocytes were inseminated with frozen-thawed boar semen and cleavage rate recorded. Before and during IVM, samples of oocytes were evaluated for LD, total and fat areas and fat gray value or for meiotic progression. Results showed that forskolin supplementation during 44 to 48 h or 22 to 24 h inhibits oocyte maturation (exp. 1: forskolin = 5.1 ± 8.0%, control = 72.6 ± 5.0%; exp. 2: forskolin = 24.3 ± 7.4%, control = 71.6 ± 5.6%) and cleavage (exp. 1: forskolin = 0.0 ± 0.0%, control = 55.4 ± 4.1%; exp. 2: forskolin = 8.3 ± 3.3%, control = 54.5 ± 3.0%). Forskolin also reduced oocyte and fat areas. In Experiment 3, forskolin negative effect on oocyte maturation and cleavage disappeared, although minor (P ⩽ 0.03) LD and oocyte fat areas were identified at 22 to 24 h of IVM. Oocytes supplemented with t10,c12 CLA during 44 to 48 h presented a lighter (P ⩽ 0.04) colour tone cytoplasm than those of control and forskolin. In conclusion, t10,c12 CLA and forskolin were capable of modifying the distribution and morphology of cytoplasmic LD during porcine oocyte maturation, thus reducing its lipid content in a time-dependent manner.

Cryopreservation of in vitro–produced sheep embryos: Effects of different protocols of lipid reduction

UNLABELLED The low survival of sheep in vitro-produced (IVP) embryos after cryopreservation is a key limiting step to the widespread of this technology. In the present work, different approaches for enhancing cryosurvival of these embryos were compared: embryo delipidation by centrifugation in the absence or presence of cytochalasin D, a cytoskeleton stabilizer or by embryo culture in the presence of different doses of the trans-10 cis-12-conjugated linoleic acid isomer (CLA). Three experiments were conducted. In experiment 1, IVP blastocysts before vitrification were randomly distributed into four groups: control; centrifuged (cent), cytochalasin D (cyto-D), centrifuged + cytochalasin D (cent + cyto-D). In experiment 2, different doses of CLA (25, 50, and 100 μM) were supplemented during embryo culture before vitrification of blastocysts. A control group ran simultaneously. A third experiment was performed to compare both approaches from the previous ones but without the groups with the worst results (groups: control, cyto-D, cent + cyto-D, CLA25, CLA50). In all experiments, embryos integrity and reexpansion were assessed after warming and after 3 hours of culture. In experiment 1, the postwarming integrity rate was the lowest (P < 0.05) in embryos from the cent group (cent: 50.6 ± 10.3% vs. CONTROL 74.6 ± 9.2%, cyto-D: 92.3 ± 9.7%, and cent + cyto-D: 90.5 ± 11.2%), whereas the best (P < 0.05) reexpansion scores were obtained in cent + cyto-D embryos (cent + cyto-D: 2.6 ± 0.28 vs. CONTROL 1.8 ± 0.08, cent: 1.9 ± 0.2, and cyto-D: 1.8 ± 0.31). In experiments 2 and 3, higher (P < 0.05) cleavage rates were observed in CLA25 (50.9 ± 6.2% and 49.2 ± 5.6%, respectively) and CLA50 (48.9 ± 6.2% and 47.6 ± 5.6%, respectively) than those in the control (41.8 ± 6.1% and 40.4 ± 5.4%, respectively) group. In experiment 2, CLA100 presented the lowest (P < 0.002) Day-6 and -7 embryo production rate and quality. After warming, superior (P < 0.02) expansion scores were achieved in CLA25 (3.1 ± 0.29) and CLA50 (3.8 ± 0.17) than in the control (1.9 ± 0.10) group. Similar results were attained in experiment 3. However, although cent + cyto-D embryos showed higher (P = 0.008) postwarming expansion scores than the control (2.8 ± 0.29 vs. 1.9 ± 0.07) group, this score was lower (P = 0.0009) than that in CLA50 embryos (3.8 ± 0.17). In conclusion, our results showed that different protocols of lipid reduction can be successfully applied to improve the cryotolerance of IVP sheep embryos.

Conservation priorities of Iberoamerican pig breeds and their ancestors based on microsatellite information.

Criollo pig breeds are descendants from pigs brought to the American continent starting with Columbus second trip in 1493. Pigs currently play a key role in social economy and community cultural identity in Latin America. The aim of this study was to establish conservation priorities among a comprehensive group of Criollo pig breeds based on a set of 24 microsatellite markers and using different criteria. Spain and Portugal pig breeds, wild boar populations of different European geographic origins and commercial pig breeds were included in the analysis as potential genetic influences in the development of Criollo pig breeds. Different methods, differing in the weight given to within- and between-breed genetic variability, were used in order to estimate the contribution of each breed to global genetic diversity. As expected, the partial contribution to total heterozygosity gave high priority to Criollo pig breeds, whereas Weitzman procedures prioritized Iberian Peninsula breeds. With the combined within- and between-breed approaches, different conservation priorities were achieved. The Core Set methodologies highly prioritized Criollo pig breeds (Cr. Boliviano, Cr. Pacifico, Cr. Cubano and Cr. Guadalupe). However, weighing the between- and within-breed components with FST and 1-FST, respectively, resulted in higher contributions of Iberian breeds. In spite of the different conservation priorities according to the methodology used, other factors in addition to genetic information also need to be considered in conservation programmes, such as the economic, cultural or historical value of the breeds involved.

Usefulness of clinical observations and blood chemistry values for predicting clinical outcomes in dairy goats with pregnancy toxaemia

BackgroundPregnancy toxaemia (PT) is a disease that affects pregnant goats during their last month of gestation and is characterized by a high case fatality rate. This study involved 32 does maintained on a commercial dairy goat farm that were diagnosed with PT. A physical examination was performed on and haematology parameters obtained from each doe, at the time of diagnosis. The data from the 24 PT goats that died was compared with the corresponding data from the 8 PT goats that survived.ResultsPolypnea, swollen limbs, anorexia with absence of ruminal motility, recumbency, nervous signs and drooping ears were the most frequently observed clinical manifestations. Nineteen out of 21 recumbent goats died. Sixteen out of 17 goats with anorexia and absence of ruminal motility died. Mean beta-hydroxybutyric acid (BHBA) values in the goats that died were not significantly different from those in goats that survived. The blood values for pH and pCO2 (p < 0.005) as well as for HCO3−, BE and K+ (p < 0.001) were significantly lower in the goats that died than in those that survived.ConclusionsThe clinical signs most indicative of a poor prognosis are anorexia with absence of ruminal motility and recumbency. Among the blood parameters to be considered, hypokalaemia and metabolic acidosis are the most relevant. Goats with PT have a high mortality and their condition can deteriorate very fast. Based on the authors’s experience, a good strategy to minimize the economic losses caused by PT is to focus on the offspring survival rate since an early decision (induction of kidding or caesarian surgery) can increase the number of alive kids.