Ultrastructural Characterization of Fresh and Vitrified In Vitro‐ and In Vivo‐Produced Sheep Embryos

The lower results in cryopreservation of in vitro‐produced (IVP) sheep embryos, when compared to the in vivo derived, limits its use. Four groups of blastocyst (BL) were evaluated: fresh IVP (n = 3), fresh in vivo derived (n = 3), warmed IVP cryopreserved in open pulled straws (OPS, n = 3) and warmed in vivo derived cryopreserved in OPS (n = 3). Ultrastructural observation of processed fresh embryos showed a reduced number of microvilli and mitochondria in the IVP ones, as well as a lower number of mature mitochondria, that can be associated with deficient metabolism in IVP embryos, possibly involved in the lower resistance to cryopreservation. Both in vivo‐derived and IVP embryos had a large number of vesicles, with light and dense content. In embryos vitrified by OPS, major changes were observed mainly in IVP embryos with small changes in grade 2 (fair) and high changes in grade 3 (bad) semithin scoring. The main changes associated with cryopreservation included disruption of cellular membranes and poor intracellular preservation, with loss of microvilli and the presence of cellular debris. In conclusion, ultrastructural evaluation of IVP blastocysts cryopreserved in OPS was herein described for the first time, reporting more severe cellular damage in these embryos when compared to those produced in vivo. This is probably associated with a lower cryotolerance that can be related to their lipid content and metabolism.     

Experimental Neospora Caninum Infection in Pregnant Dairy Heifers Raises Concentrations of Pregnancy‐Associated Glycoproteins 1 and 2 in Foetal Fluids

Plasma concentrations of PAG‐1 are used for pregnancy diagnosis and as a marker of placental/foetal well‐being, while those of PAG‐2 may be an indicator of abortion risk in Neospora caninum‐infected cows. Studies have shown that N. caninum infection modifies PAG‐1 and PAG‐2 patterns in maternal blood plasma. However, no prior work has examined the effects of N. caninum infection on concentrations of PAGs in foetal fluids. In this study, PAG‐1, PAG‐2 and pH levels were determined in the amniotic and allantoic fluids of foetuses collected at 152 days of gestation from control uninfected dams and from dams experimentally infected with N. caninum on Day 110 of gestation. Foetal fluids from infected foetuses had significantly higher PAG‐2 concentrations (p = 0.026) and pH values (p = 0.02) than fluids from non‐infected foetuses. In infected foetuses, significantly higher concentrations of PAG‐1 (p < 0.001) and PAG‐2 (p < 0.001) were detected in fluid samples showing antibodies against N. caninum than those without antibodies. Moreover, pH values were significantly higher (p = 0.011) in foetal fluid samples with antibodies than in samples from non‐infected foetuses. In conclusion, this is the first report on the effect of N. caninum infection on PAG levels in foetal fluids. Our results indicate that following the experimental infection of dams with N. caninum on Day 110 of gestation, foetal fluids collected from the infected foetuses of these dams featured higher PAG‐1 and PAG‐2 levels and pH values than fluids from non‐infected controls, provided that the samples tested showed the presence of antibodies. The clinical implications of these findings are that following infection with N. caninum, most cows will experience some level of placental damage and that this injury correlates with foetal fluid PAG levels and pH.     

The effect of induced forelimb lameness on thoracolumbar kinematics during treadmill locomotion

REASONS FOR PERFORMING STUDY: Lameness has often been suggested to result in altered movement of the back, but there are no detailed studies describing such a relationship in quantitative terms. OBJECTIVES: To quantify the effect of induced subtle forelimb lameness on thoracolumbar kinematics in the horse. METHODS: Kinematics of 6 riding horses was measured at walk and at trot on a treadmill before and after the induction of reversible forelimb lameness grade 2 (AAEP scale 1-5). Ground reaction forces (GRF) for individual limbs were calculated from kinematics. RESULTS: The horses significantly unloaded the painful limb by 11.5% at trot, while unloading at walk was not significant. The overall flexion-extension range of back motion decreased on average by 0.2 degrees at walk and increased by 3.3 degrees at trot (P<0.05). Changes in angular motion patterns of vertebral joints were noted only at trot, with an increase in flexion of 0.9 degrees at T10 (i.e. angle between T6, T10 and T13) during the stance phase of the sound diagonal and an increase in extension of the thoracolumbar area during stance of the lame diagonal (0.7degrees at T13, 0.8 degres at T17, 0.5 degres at L1, 0.4 degrees at L3 and 0.3 degrees at L5) (P<0.05). Lameness further caused a lateral bending of the cranial thoracic vertebral column towards the lame side (1.3 degrees at T10 and 0.9 degrees at T13) (P<0.05) during stance of the lame diagonal. CONCLUSIONS: Both range of motion and vertebral angular motion patterns are affected by subtle forelimb lameness. At walk, the effect is minimal, at trot the horses increased the vertebral range of motion and changed the pattern of thoracolumbar motion in the sagittal and horizontal planes, presumably in an attempt to move the centre of gravity away from the lame side and reduce the force on the affected limb. POTENTIAL RELEVANCE: Subtle forelimb lameness affects thoracolumbar kinematics. Future studies should aim at elucidating whether the altered movement patterns lead to back and/or neck dysfunction in the case of chronic lameness.     

Chronological study of Mycoplasma hyopneumoniae infection, seroconversion and associated lung lesions in vaccinated and non-vaccinated pigs

A field trial was conducted to study Mycoplasma hyopneumoniae (Mh) infection dynamics by nested polymerase chain reaction (nPCR) and serology in pigs of a farm affected by enzootic pneumonia (EP). Moreover, correlation of Mh detection at different respiratory tract sites with presence of EP gross and microscopic lung lesions was assessed. These parameters were studied and compared between vaccinated (two doses at 1 and 3 weeks of age versus one dose at 6 weeks of age) and non-vaccinated pigs. Animals were monitored from birth to slaughter by nPCR from nasal swabs and by serology. From 3 to 22 weeks of age, an average of three pigs per treatment and per batch were necropsied (n = 302). The remaining pigs were sent to the slaughter (n = 103). Nasal, bronchial and tonsillar swabs were taken from the necropsied/slaughtered pigs; gross and microscopic EP-suggestive lung lesions were also assessed. Single and double vaccination resulted in earlier seroconversion and higher percentage of Mh seropositive pigs compared to control group. At slaughter, double vaccinated pigs showed lower percentage of EP-compatible gross lung lesions and lower Mh prevalence at upper respiratory tract sites (nasal cavity and tonsil) than control pigs.     

Phenotypic and genotypic characterization of Paenibacillus larvae isolates

Paenibacillus larvae is the causative agent of American Foulbrood (AFB), a severe disease of honeybees (Apis melifera). The aim of this work was to develop a strategy for the subtyping and the epidemiological analysis of P. larvae. Phenotypic characterisation, susceptibility to several antibiotics, electrophoresis of whole bacterial proteins, rep-PCR, ribotyping and DGGE were assessed using a collection of P. larvae isolates from different Uruguayan and Argentinean locations. Results indicated that there are two P. larvae genotypes circulating in Uruguay ERIC I-BOX A (worldwide distributed) and ERIC I-BOX C (exclusively detected in Argentina until this study). These results suggest that P. larvae isolates had moved between Argentina and Uruguay, probably through the Uruguay River. Patterns of whole bacterial proteins, DGGE and ribotyping did not improve the P. larvae intraspecific discrimination. Antibiotic susceptibility assays showed that 100% isolates were OTC-sensitive and 22% (belonging to ERIC I-BOX A group) were sulfisoxazole-resistant. This work may contribute to the elucidation of basic aspects related to the epidemiology of AFB in Uruguay and in the region.