Fernando Caetano de Oliveira

Identificação de Neospora caninum por PCR em cordão umbilical de bezerros recém nascido no Brasil

This study was conducted at a beef cattle breeding farm in the far southern region of Brazil. The birth of a calf with unilateral corneal opacity was immediately reported to the Laboratory of Parasitology, in the Universidade Federal de Pelotas, Rio Grande do Sul, Brazil; blood was collected from the cow and calf before colostrum intake. The umbilical cords from this calf and from six other healthy animals were collected. Serological examination, utilizing an indirect fluorescent antibody test, was done using a cut-off point of 1:100. Polymerase chain reaction (PCR) was also performed using the umbilical cord samples. Serological tests showed that the calf was positive for Neospora caninum at birth, with a titer of 1:1600; a titer of 1:3200 was reported in the dam. PCR, using umbilical cord tissue from the affected animal, was positive for the presence of this parasite, and the molecular identity of the amplified product was confirmed by sequencing. Therefore, the detection of N. caninum DNA in the umbilical cord represents a novel alternative test for the diagnosis of this parasitic infection in newborn calves that are clinically suspected to have neosporosis.

Expression of growth and differentiation Factor 9 and cognate receptors during final follicular growth in cattle

Mutations in growth and differentiation factor 9 (GDF9) gene are associated to sterility or, paradoxically, increased ovulation rate in ewes. Despite its importance, the exact function of GDF9 in ovarian physiology is still poorly understood. This study aimed to investigate GDF9 function during dominant follicle growth and its regulation in follicular fluid. The regulation of GDF9 receptors in GnRH/LH-stimulated granulosa cells was also investigated. In a first experiment, a new follicular wave was induced and the intrafollicular GDF9 treatment into the largest growing follicle (8.5-9.5 mm) at both 100 (n = 3) and 1000ng/ml (n = 4) had no effect on follicular growth, estrus manifestation and ovulation compared to control (PBSinjected) follicles (n = 3). In a second experiment, follicles were obtained just after follicular deviation (day 4 after follicular emergence) and the abundance of GDF9 in follicular fluid did not differ between healthy dominant (n = 4) and atretic subordinate follicles (n = 4), as assessed by western blot analysis. Finally, mRNA expression of BMPR2 and TGFBR1 receptors was evaluated in granulosa cells obtained from preovulatory follicles (>12 mm diameter) obtained 0, 3, 6, 12 or 24 h after i.m. GnRH administration (n = 4-5 follicles/moment). Both receptors were significantly up regulated 12 h after GnRH treatment. Present results do not confirm the hypothesis that GDF9 inhibits dominant follicle growth and suggests a minor role in determining follicle fate. In the other hand, GDF9 receptors regulation in GnRH/LH-stimulated granulosa cells provides the first in vivo evidence of its involvement in the complex cascade of events that culminates in ovulation and luteinization in cattle.

Motilidade de espermatozoides bovinos após incubação em fluído folicular

The objective of this work was to evaluate sperm cell motility after intrafollicular artificial insemination (IFAI) in vivo or after incubation in follicular fluid in vitro. In the in vivo experiment, IFAI was performed, followed by the recovery of follicular content 1 to 4 hours later, in order to assess sperm motility. In the in vitro experiment, spermatozoa from a pool of commercial frozen-thawed semen were evaluated for their kinetics after incubation for 1 or 3 hours, either pure (pool, control group) or in follicular fluid (FF). A low motility of sperm cells was observed in the FF samples, both in vitro and in vivo. In vitro, the main parameters negatively affected in the sperm cells incubated in FF, compared with the control, were: total motility (TM), progressive motility (PM), curvilinear distance, and straightness, after 1 hour of incubation; and TM, PM, average path velocity, and curvilinear velocity after 3 hours of incubation. The ovarian follicle and follicular fluid do not provide a suitable environment to maintain bovine sperm cell motility.