Toll-like receptor and related cytokine mRNA expression in bovine corpora lutea during the oestrous cycle and pregnancy.

Improving our understanding of the mechanisms controlling the corpus luteum (CL) and its role in regulating the reproductive cycle should lead to improvements in the sustainability of today's global animal industry. The corpus luteum (CL) is a transient endocrine organ composed of a heterogeneous mixture steroidogenic, endothelial and immune cells, and it is becoming clear that immune mechanisms play a key role in CL regulation especially in luteolysis. Toll-like receptors (TLR) mediate innate immune mechanisms via the production of pro-inflammatory cytokines, especially within various tissues, although the role of TLR within CL remains unknown. Thus, the objectives of this study were to characterize TLR mRNA expression in the CL during the oestrous cycle and in pregnancy (day 30-50), and to examine the role of TLR signalling in luteal cells. Corpora lutea were collected at various stages of the cycle and pregnancy and analysed for TLR and cytokine mRNA expression. In addition, luteal cells were cultured with the TLR4 ligand (lipopolysaccharide, LPS) for 24 h to evaluate the role of TLR4 in regulating luteal function. Toll-like receptors 1, 2, 4, 6, tumour necrosis factor alpha (TNF), interferon gamma (IFN-G), and interleukin (IL)-12, mRNA expressions were greatest in regressing CL compared with earlier stages (p < .05), whereas no change was observed for IL-6 mRNA expression. Cytokine mRNA expression in cultured luteal cells was not altered by LPS. Based on these data, one or more of the TLRs found within the CL may play a role in luteolysis, perhaps via pro-inflammatory cytokine mRNA expression.

Pregnancy recognition and abnormal offspring syndrome in cattle.

Development of the post-hatching conceptus in ruminants involves a period of morphological expansion that is driven by complex interactions between the conceptus and its intrauterine environment. As a result of these interactions, endometrial physiology is altered, leading to establishment of the pregnancy and continued development of the placenta. Disruption of normal fetal and placental development can occur when embryos are exposed to manipulations in vitro or when inappropriate endocrine sequencing occurs in vivo during the pre- and peri-implantation periods. The present review addresses the development of the post-hatching bovine conceptus, its interactions with the maternal system and changes in development that can occur as a result of in vivo and in vitro manipulations of the bovine embryo.

Maternal serum progesterone concentration and early conceptus development of bovine embryos produced in vivo or in vitro.

The hormone progesterone is essential for proper embryonic development. The objective of this study was to examine the relationship between recipient serum concentrations of progesterone, at the time of embryo transfer and at conceptus recovery, on conceptus development from in vivo- or in vitro-produced embryos. Embryos were produced in vivo by superovulation of Holstein cows (IVO; n = 17) or in vitro with either serum-containing (IVPS; n = 27) or serum-restricted medium (IVPSR; n = 34). Single grade I blastocysts from each embryo production system were transferred into heifers on day 7 of development. Conceptuses were recovered on day 17 of gestation and classified as complete, degenerated, or no conceptus. Compared with the IVO group, in vitro-produced embryos had more (P = 0.055) degenerated conceptuses (IVO, 0%; IVPS, 18.5%; and IVPSR, 20.6%). There were no differences in progesterone concentrations at the time of transfer when recipients received either male or female embryos (P > 0.05). Progesterone concentrations in recipients receiving in vivo-produced embryos were higher (P < 0.05; 3.74 ± 0.4 ng/mL; least-squares mean ± standard error of the mean) on day 7 compared with those receiving in vitro-produced embryos (IVPS, 2.4 ± 0.2; IVPSR, 2.58 ± 0.3 ng/mL). However, there was no difference in progesterone concentration on day 7 between treatment groups for heifers from which short conceptuses (≤194 mm) were recovered on day 17. In contrast, when longer (>194 mm) conceptuses were recovered on day 17, heifers receiving in vitro-produced embryos had lower (P = 0.05) serum concentrations of progesterone on day 7 compared with those receiving in vivo-produced embryos (IVPS, 2.2 ± 0.5; IVPSR, 2.3 ± 0.5; IVO, 3.9 ± 0.5 ng/mL). In conclusion, differences in autonomy may exist between in vitro- and in vivo-produced embryos during the period of conceptus elongation with in vitro-produced embryos relying more on intrinsic factors to influence elongation.

Detection of mRNA for inhibin alpha- and beta A-subunits in bovine ovarian tissues and the effect of in vivo administration of GNRH.

The aims of these studies were to determine which types of bovine ovarian tissue contain mRNA for inhibin/activin subunits and whether administration of GnRH influences concentration of these mRNAs. In experiment (exp.) one, cows in the luteal phase of the estrous cycle were given prostaglandin F2 alpha (PGF2 alpha) to induce luteal regression and injected after 40 hr with saline (n = 5) or 100 micrograms GnRH (n = 6). Ovaries were removed 6 hr later. In exp. two, unilaterally ovariectomized (OVX) heifers (n = 33) in the luteal phase of their estrous cycle were given PGF2 alpha to induce luteal regression. Twelve heifers were OVX without injection of GnRH at 24 (n = 6) or 40 hr (n = 6) after PGF2 alpha. The remaining heifers (n = 21) were given 100 micrograms GnRH at 40 hr after PGF2 alpha injection and OVX 8 (n = 4), 16 (n = 5), 24 (n = 6) or 48 (n = 6) hr after GnRH injection. Total cellular RNA was isolated from large follicles (exp. one and two), small-medium follicles and stromal tissue (SMS) and corpora lutea (CL; exp. one) tissues and analyzed by dot blot and Northern blot techniques by hybridizing with cDNA probes for human inhibin/activin alpha- and beta A-subunits. Large follicles were classified as steroidogenically active (EA) if follicular fluid (FF) concentration of estradiol-17 beta (E2) was greater than progesterone (P4), or if P4 and E2 concentrations in FF were greater than 100 ng/ml, and estrogen inactive (EI) if FF concentration of E2 and P4 did not satisfy these criteria. In exp. one, mRNA for the alpha-subunit was primarily expressed in EA follicles, and detectable in EI follicles, SMS, and CL while beta A-subunit mRNA was detected only in large EA follicles and a few SMS samples. The mRNA (x +/- SEM fmoles/mg DNA) for both subunits of inhibin/activin was higher (P < .05) in EA follicles from GnRH-treated cows (alpha = 210.2 +/- 38.6; beta A = 376.9 +/- 41.0) than in EA follicles from control cows (alpha = 102.5 +/- 28.6; beta A = 170.8 +/- 57.6). Concentration of mRNA for the alpha-subunit of inhibin in other ovarian tissues was not different (P > .10) between saline and GnRH treatments.(ABSTRACT TRUNCATED AT 400 WORDS)

Birth of a Holstein freemartin calf co-twinned to a schistosomus reflexus fetus.

An unusual case of a live Holstein freemartin calf co-twinned with schistosomus reflexus fetus is presented here. Delivery of the schistosomus reflexus was achieved by fetotomy 24 h after manual delivery of a live heifer calf. The dam subsequently experienced concurrent metritis and left displacement of the abomasum; however, she conceived following insemination 85 d post partum. Cytogenetic examination of the live heifer using lymphocyte culture and cutaneous fibroblast cell culture failed to demonstrate chromosomal chimerism, whereas polymerase chain reaction (PCR) detected the presence of the bovine Y-chromosome marker BRY-1. Low concentrations of testosterone, estradiol and progesterone at 3, 6, 24 and 48 h after administration of hCG were detected in the serum of the freemartin heifer. Genetic, hormonal, histological and clinical findings established the live female co-twin calf was a freemartin, an abnormality of phenotypic sex. Failure to detect any significant peripheral concentrations of androgen supports the hypothesis that masculinization of the freemartin reproductive tract arises from diffusion of androgen and possibly other substances from the male co-twin rather than from endogenous production of androgen within the freemartin. This report documents that the freemartin condition can be induced by a male fetus with severe developmental abnormalities.

Influence of in vitro systems on embryo survival and fetal development in cattle.

In vitro systems are commonly used for the production of bovine embryos. Comparisons between in vivo and in vitro produced embryos illustrate that the morphology of preimplantation-stage embryos differ significantly, the survival of embryos and fetuses is decreased, the size distributions of the populations of conceptuses and fetuses are altered throughout gestation, and placental development is significantly changed. Taken together these findings indicate that exposure to some in vitro environments during the first 7 days of life can profoundly influence fetal and placental development in cattle. An understanding of how in vitro oocyte maturation, in vitro fertilization, and embryo culture systems influence both fetal and placental development should result in systems that consistently produce normal embryos, fetuses, and calves.

Estimates of pregnancy outcomes based on selection of bovine embryos produced in vivo or in vitro.

The objective of this study was to estimate the degree of variation among experienced evaluators selecting in vivo- or in vitro-produced embryos for transfer and to determine how this affects both the proportion of recipients becoming pregnant after transfer, and the number of embryo transfers required per pregnancy. Data from 6 experienced evaluators who graded Day 7 embryos produced either in vivo (n = 15) or in vitro (n = 15) were used to estimate these effects. The evaluators viewed video recorded images of the embryos and classified each embryo for stage of development and quality grade (1 = excellent, 2 = good, 3 = fair, 4 = degenerated and nontransferable). The statistical model considered transfer of embryos of the following individual or combined grades: Grade 1 only, Grade 2 only, Grade 3 only, Grades 1 and 2, Grades 2 and 3, and Grades 1, 2 and 3. Probabilities of pregnancy after embryo transfer were based on pregnancy rates of recipients at the facility of 1 of the 6 evaluators where the percentages of heifers pregnant after the transfer of Grade 1, 2 and 3 embryos, by embryo source, were 76, 65 and 54% (in vivo), and 59, 45 and 30% (in vitro). Within most grades, the proportion of embryos selected for transfer differed (P < 0.05) among the 6 evaluators. Although no significant differences (P > 0.10) were found among evaluators in the proportion of recipients pregnant after transfer within any embryo grade, there was substantial variation among evaluators in the proportion of recipients becoming pregnant, especially for embryos produced in vitro. Estimated percentages of heifers becoming pregnant for embryos classified as Grade 1, 2 or 3 were 66 to 76, 62 to 69, and 54 to 60%, respectively, for in vivo-produced embryos; and, 39 to 59, 15 to 45, and 24 to 32%, respectively, for in vitro-produced embryos. Approximately twice as many transfers were needed per pregnancy for embryos produced in vitro as for those produced in vivo regardless of the grade.

Agreement among evaluators of bovine embryos produced in vivo or in vitro.

Six experienced individuals evaluated 40 embryos on videotape for stage of development and quality grade. These 40 observations comprised 15 embryos produced in vivo, 15 embryos produced in vitro, and 10 embryos that were repeated throughout the videotape. Embryos produced in vivo were recovered from uterine flushings of superovulated heifers 7 d after estrus, and embryos produced in vitro were harvested 7 d after insemination of in vitro-matured oocytes. Embryos of various stages (morulae, blastocysts, or degenerated) and quality grades (1 = excellent, 2 = good, 3 = fair, 4 = degenerated) were recorded on videotape for evaluation. After video microscopy, the embryos were stained and the number of nuclei per embryo was counted. Six evaluators reviewed the videotape and the percentage of agreement and kappa (k; agreement beyond chance) among evaluators were determined for classifications of stage and grade. Consistency of each evaluator's responses was estimated using the 10 repeated embryos. Agreement within evaluators was higher for stage of embryo development (89.2%) than quality grade (68.5%). Agreement among evaluators for stage was slightly higher with embryos produced in vivo (85.0%, k = 0.74) than in vitro (72.3%, k = 0.48). Agreement among evaluators for grade was similar with embryos from in vivo (61.0%, k = 0.46) and in vitro (57.7%, k = 0.42) production. For both sources of embryos, agreement was substantially better for Grades 1 and 4 than for Grades 2 and 3. The results of this study suggest that good to excellent agreement exists for classifying Day 7 bovine embryos by stage and by extremes of quality grade (Grades 1 and 4) but not by degree of abnormal morphology (Grades 2 and 3). Simple grading criteria of Grade 1 (highest quality), Grade 2 (morphologic defects), and Grade 3 (degenerated) maximized agreement among evaluators.

Relationships between breeding soundness and sex drive classifications in beef bulls.

Prior to use in single-sire mating trials, ninety-two beef bulls were evaluated to determine the relationships between. Breeding Soundness Examination (BSE) and sex drive assessment categories. A BSE score was given to each bull based on scrotal circumference, spermatozoal motility and morphology. Eighty bulls were classified as satisfactory potential breeders, while 12 were classified as questionable potential breeders. Each bull was exposed to two, 10 min modified libido/serving capacity tests. Sixty-nine bulls were classified as high libido and 23 were classified as medium libido. Differences in scrotal circumference and spermatozoal motility did not influence (P > 0.05) the BSE classifications. However, differences in primary, secondary and total spermatozoal abnormalities were the major factors affecting the BSE classification. Bulls classified as high libido serviced three times more (P < 0.01) during the two test periods than bulls classified as medium libido. High libido bulls were superior in all sex drive traits. However, bulls in the two libido categories did not differ (P > 0.05) in semen quality or scrotal circumference. There was no relationship (P > 0.05) between BSE and sex drive traits (r = -0.16 to 0.24).

Beef bulls mated to estrus synchronized heifers: Single- vs multi-sire breeding groups.

Beef bulls of approximately 15 months of age were placed with heifers at bull to female ratios (BFR) of 1:20 (n = 3) and 2:40 (n = 3) to compare the breeding efficiency of bulls used in either a single- or a multi-sire group. Prior to the breeding period, each bull was given a breeding soundness exam and two exposures to a libido/serving capacity test. For the purpose of synchronizing estrus, heifers received a nine-day Syncro-Mate-B((R)) (G. D. Searle & Co.) treatment. Twenty-seven hr after removal of the implants, bulls were placed with heifers and sexual activity was observed continuously for the succeeding 30 hr. With the exception of number of services per heifer, the mating performance of bulls and pregnancy rates at both BFR's were not different (P>.05). Heifers in single-sire groups were serviced more times (P<.05) than those in multi-sire groups (4.1 +/- 0.6 vs 2.6 +/- 0.2, respectively). Approximately 50% of heifers in multi-sire groups were serviced by both bulls. Due to the overlap in servicing heifers and the non-significant difference in pregnancy rates between the two BFR's, it was concluded that a single-sire mating program was more efficient.

Serum progesterone concentration in pyometritic and normal postpartum dairy cows.

Two studies were conducted to evaluate the concentration of serum progesterone in pyometritic cows and relate it to palpation of ovarian structures per rectum . In Trial 1, serum samples from 34 pyometritic cows were assayed for progesterone. Mean serum progesterone concentrations were 6.8 +/- 0.7 ng/ml. In Trial 2, each of 54 pyometritic cows was paired with a control cow on the basis of days post partum (18-50 days). Mean concentration of progesterone was 9.7 +/- 1.0 ng/ml for the pyometritic cows and 5.7 +/- 0.8 ng/ml in control cows (P<0.005). Progesterone concentration was greater (P<0.005) in both groups of cows with palpable corpora lutea (CLs). Ninety-six percent of the pyometritic cows had palpable CLs compared to 57% of the control group. Comparing serum progesterone only in cows with a palpable CL, the mean concentration was still greater (P<0.005) in the pyometritic group (10.6 +/- 1.0 ng/ml) than the control group (6.6 +/- 1.0 ng/ml). Compatability of rectal palpation findings and concentrations of serum progesterone were 92.6% for the pyometritic group and 72.2% for the control group. Progesterone concentration increased (P<0.05) by increased days post partum in Trial 2 (n=54) but not in Trial 1 (n=23). In both Trials 1 and 2, uterine size due to pyometra increased (P<0.05) with increased days post partum. No other associations were found.

Breeding soundness and sex drive by breed and age in beef bulls used for natural mating.

A total of 92 range beef bulls (Hereford = 60; Angus = 32) were given a breeding soundness examination (BSE) and two assessments for sex drive prior to their use in 23 breeding trials employing estrous synchronized females. Bulls were in three age groups: yearlings (n=29), two year olds (n=36), and three year olds and older (n=27). All yearling bulls were virgins, but the majority of the older bulls had previous mating experience. Angus bulls were superior (P<0.01) to Herefords in spermatozoal morphology and BSE score. Scrotal circumference increased with age beyond two years in Angus bulls but not in Herefords. Spermatozoal abnormalities generally decreased with age. BSE scores did not differ significantly among age groups. Apart from number of mounts, measures of sex drive did not differ with age or breed of bulls. This represents qualified justification for the current practice of using the same sex-drive assessment procedures for Bos taurus bulls of various ages and breeds.

Diagnosis of luteal and follicular ovarian cysts in dairy cows by sector scan ultrasonography.

Sixty-seven ultrasonograms of ovarian cysts (cysts) from 35 cows were used to evaluate sector scan ultrasonography as a means for differentiating luteal cysts from follicular cysts. Initial diagnosis of cysts was made by ovarian palpation per rectum during weekly herd visits. The ovaries of each cow were then examined by ultrasonography. Ultrasonograms of cysts > 25 mm in diameter were diagnosed as either luteinized or follicular cysts and were recorded on video tape for evaluation by a second clinician. Serum progesterone concentrations at the time of examination were determined by radioimmunoassay and were used to classify luteal (> 0.5 ng/ml) or follicular (

Breeding soundness, libido and performance of beef bulls mated to estrus synchronized females.

Ninety-three beef bulls and 2316 females were used to determine the relationships between breeding assessments of bulls and subsequent mating performance. Each bull was given a breeding soundness examination (BSE) and two 10-min libido/serving capacity (L/SC) tests. Breeding potential of each bull was classified as satisfactory (BSE score = 60 to 100) or questionable (BSE score = 30 to 59); libido was classified as either high (mean score = 9.0 to 10) or medium (mean score = 6.0 to 8.5). Bulls were then joined, single-sire, with groups of females which had received one of two treatments to synchronize estrus. Bull-to-female ratios ranged from 1:7 to 1:51. Continuous observations were conducted on the mating activity of each group. One bull was removed from the experiment due to a severe breeding problem. Bulls of satisfactory breeding potential (n = 80) did not differ (P > 0.10) from bulls of questionable breeding potential (n = 12) in measurements of mating activity. However, by the end of the synchronized breeding period, bulls classified as satisfactory breeders achieved approximately a 9% higher (P < 0.10) pregnancy rate than did bulls of questionable breeding status (45.6 +/- 2.1% vx 36.5 +/- 5.3%). Bulls with a high libido (n = 69) serviced more (P < 0.01) estrous females (81.3 +/- 3.1% vs 63.5 +/- 4.2%) than bulls with a medium libido (n = 23). However, pregnancy rates achieved by bulls of either libido classification did not differ significantly. Individual components of the BSE as well as mean libido score were poorly correlated with pregnancy rates (r = -0.22 to 0.18). It was concluded that classification of bulls by mean libido score can aid in identifying groups of bulls that service more estrus synchronized females, whereas classification by BSE score aids in identifying groups of bulls that impregnate more females.

Ovarian follicular development following administration of progesterone or aspiration of ovarian follicles in Holstein cows.

The objective of this study was to compare the effects of administration of a single injection of progesterone (P4) and follicle aspiration on Day 7 of the estrous cycle on the timing and synchrony of follicular wave emergence, time of ovulation, and concentrations of P4, estradiol and FSH in Holstein cows. Twenty cows were assigned to 4 groups (n=5 cows per group) in a 2 by 2 factorial arrangement. Cows were treated on Day 7 (Day 0 = estrus) of the estrous cycle with either sham follicular aspiration and an oil vehicle administered intramuscularly (control), aspiration of ovarian follicles (aspiration), 200 mg of P4 im, or aspiration and 200 mg of P4 im (aspiration + P4). On Day 11, PGF(2alpha)(25mg) was administered to all groups. Synchrony of ovulation was less variable in each of the treatment groups compared with the control group (P<0.05), whereas ovulation was delayed in cows in the P4 group (P<0.05). Day of follicular wave emergence was delayed in the cows of the P4 group compared with cows in the aspiration and aspiration + P4 groups (P<0.01), whereas variability in wave emergence was less among both groups of aspirated cows compared with the cows in the control group (P<0.01). More follicles 4 to 7 mm in diameter were detected in the 2 aspiration groups compared with the cows in the control and P4 group (P<0.05). No difference was detected among groups in the maximum concentration of FSH associated with follicular wave emergence. We conclude that both the administration of P4 and the aspiration of follicles on Day 7 of the estrous cycle improves the synchrony of ovulation when luteolysis is induced on Day 11 and results in similar concentrations of FSH at the time of follicular wave emergence, but the timing of wave emergence and the number of follicles post-emergence differ.

Effect of Actinomyces pyogenes and gram-negative anaerobic bacteria on the development of bovine pyometra.

Fifteen lactating Holstein cows were used in a trial designed to evaluate the effectiveness of intrauterine inoculation (challenge) of Actinomyces pyogenes (A) alone or in combination with Fusobacterium necrophorum (F) and Bacteroides melaninogenicus (B) to induce pyometra. Cows were assigned to one of five groups: A (n = 3), AB (n = 3), AF (n = 3), ABF (n = 3) or C (control, broth medium alone; n = 3). All cows exhibited estrus 12 or 13 d prior to challenge (Day 0=first day of challenge). During the prechallenge period, the reproductive tract of each cow was palpated per rectum and uterine fluid aspirates for culture and uterine biopsies were also obtained. All cows received an intravenous injection of 5,000 IU of human chorionic gonadotropin (hCG; Day 5) and an intrauterine infusion of 40 ml of 0.7% iodine solution (Day 1). Cows were then inoculated on Days 0, 1 and 2 of the experiment. Sequential palpations of the reproductive tracts, samples of uterine fluid for culture and uterine biopsies were performed for a total of 30 d after the first inoculation. A cow was diagnosed as having pyometra when purulent uterine fluid and a corpus luteum were detected by palpation per rectum. The number of cows that developed pyometra in Group A was 2 of 3, in Group AB 3 of 3, in Group AF 3 of 3, in Group ABF 3 of 3 and in Group C 0 of 3. Cows with pyometra did not exhibit estrus. In 7 of 11 cows, pyometra persisted for more than 21 d. In cows that developed pyometra, the same species of bacteria infused into the uterus were usually recovered one or more times during the postchallenge period. When clinical pyometra was diagnosed, histologic endometritis was invariably present. Histologic endometritis and concurrent isolation of A . pyogenes alone or A . pyogenes with gram-negative anaerobic bacteria occurred in 91.7% of samples during the postchallenge period. Regardless of bacterial treatment, gram-negative anaerobic bacteria were frequently isolated with A . pyogenes during this period.

Efficiency of single-sire mating programs with beef bulls mated to estrus synchronized females.

Efficiency in reference to pregnancy rates of breeding beef bulls with estrus synchronized cows and heifers was tested. Most bulls (104 of 112) were given a breeding soundness examination and two 10-min libido/serving capacity tests. Females received either Syncro-Mate-B (SMB) or two injections of Prostaglandin F(2)alpha (PGF) to synchronize estrus. They were assigned to single-sire breeding groups with bull-to-female ratios ranging from 1:7 to 1:51. Control groups consisted of untreated females maintained in single-sire breeding pastures with ratios from 1:24 to 1:37. Continuous observations of sexual activity were made for 30 h (SMB) and 48 h (PGF). After the 120-h posttreatment breeding period, females were placed in breeding pastures. During the synchronized breeding period the percentage of pregnant cattle of total treated was 43.5 +/- 1.7% compared (P < 0.01) with 58.9 +/- 3.3% for the control group after 23 d of breeding. At end of 28-d (treated) and 46-d (control) period, the percentage of pregnant females was 75.0 +/- 2.4 and 79.6 +/- 4.7, respectively (P > 0.05). In SMB trials, the percentages of females exhibiting estrus, those serviced at estrus and those pregnant following service during the synchronized breeding period were 90.8 +/- 1.5, 73.3 +/- 4.5 and 56.4 +/- 5.6%, respectively. In PGF trials, the means for these same factors were 78.3 +/- 2.4, 70.4 +/- 5.9 and 56.1 +/- 6.5%, respectively.

Aspects of bacteriology and endocrinology of cows with pyometra and retained fetal membranes.

In an effort to clarify the pathogenesis of pyometra, 20 cows with retained fetal membranes and 20 without, but with contemporary calving dates were studied. They were palpated and their uteri were subjected to sample collections for bacteriologic cultural examinations twice weekly for 4 weeks. Blood samples were obtained each day and evaluated for serum progesterone concentration. Three cows without and 3 with retained fetal membranes developed pyometra during the study, resulting in 3 groups designated control (CON), cows with retained fetal membranes (RFM), and cows with pyometra (PYO). Bacterial isolations occurred less frequently in the CON group than in the PYO and RFM groups. Growth patterns of bacteria also varied between groups. Coliform and incidental bacteria disappeared from the uterus of the PYO group by the end of the 3rd week. In contrast, heavy growth of Corynebacterium pyogenes and gram-negative anaerobic bacteria developed during this same period in the PYO group. In cows with pyometra, the significant persistent pathogenic bacteria recovered were C pyogenes and gram-negative anaerobic bacteria, especially Fusobacterium necrophorum and Bacteroides melaninogenicus. Anaerobic bacteria were isolated simultaneously with C pyogenes in most cows of the PYO group, but less often in CON and RFM groups, and highest growth levels were present near the time of ovulation. Clinically, pyometra usually developed about 10 days after observation of concurrent ovulation and high growth levels of C pyogenes and gram-negative anaerobic bacteria. A hypothesis is presented for development of pyometra in the cow.

Diagnosis of luteal and follicular ovarian cysts by palpation per rectum and linear-array ultrasonography in dairy cows.

The purpose of this study was to determine and compare the accuracy of palpation per rectum and linear-array ultrasonography for diagnosing follicular vs luteal ovarian cysts in cows. Forty-seven examinations of ovarian cysts from 28 cows were diagnosed by palpation per rectum as either a firm, thick-walled structure (luteal cyst) or a soft, thin-walled structure (follicular cyst) during weekly herd examinations. The ovaries of each cow were then examined by ultrasonography. Ultrasonograms of cysts greater than 25 mm in diameter were diagnosed as luteal or follicular cysts and were recorded on videotape for evaluation by a second clinician. Serum progesterone concentrations at the time of examination were determined by radioimmunoassay and used to classify luteal (greater than 0.5 ng/ml) or follicular (less than or equal to 0.5 ng/ml) cysts. Selection of this discriminatory level was based on response of a proportion of cows with luteal cysts that were given 25 mg of prostaglandin F2 alpha at the time of diagnosis by ultrasonography. Sensitivity and specificity of palpation per rectum for diagnosis of type of ovarian cyst were low (43.3 and 64.7%, respectively). In contrast, sensitivity and specificity of ultrasonography were considerably higher (86.7 and 82.3%, respectively). Agreement between the 2 methods of diagnosis was 57.4%. Overall agreement between the 2 clinicians' diagnoses by ultrasonography was 85.1%. On the basis of our findings, we confirm that luteal and follicular cysts cannot be accurately differentiated by palpation per rectum alone. These data suggest that linear-array ultrasonography is more effective than palpation per rectum for diagnosing type of ovarian cyst in cows.

Antimicrobial therapy for rams with Brucella ovis infection of the urogenital tract.

Rams shedding Brucella ovis in semen but without palpable abnormalities of the epididymides were treated with long-acting oxytetracycline for 15 days and dihydrostreptomycin for 7 days (n = 9) or conventional oxytetracycline and dihydrostreptomycin (n = 9) for 7 days. Nine rams were not treated. More treated rams were considered to have satisfactory breeding soundness examination results at posttreatment weeks 3, 7, 12, and 19. Nontreated rams continued to shed B ovis in semen. After treatment, B ovis was not recovered from 78% of rams given long-acting oxytetracycline and dihydrostreptomycin or from 89% of rams given conventional oxytetracycline and dihydrostreptomycin. At week 21, all rams were euthanatized, and specimens of the testes and epididymides were bacteriologically cultured for B ovis. Brucella ovis was not recovered from the testes of rams or from the epididymides from rams not shedding the organism in the semen. In one treated ram, B ovis was recovered from the semen but not from other tissues. All rams remained ELISA-positive, with the exception of 2 treated rams that ceased shedding B ovis in semen immediately after treatment was started; both these rams became ELISA-negative on the last examination at week 19.