Takeshi Osawa

Plasma anti-Müllerian hormone as a biomarker for bovine granulosa-theca cell tumors: Comparison with immunoreactive inhibin and ovarian steroid concentrations

Granulosa-theca cell tumors (GTCTs) are the most frequently reported ovarian tumors in cattle. Clinically, GTCTs could be confused with other ovarian abnormalities; therefore, the only definitive diagnosis for such tumors is histopathology of a biopsy from the affected ovary. However, this is an invasive technique and unsuitable for farm conditions. As a result, the key aim of this study was to evaluate the diagnostic value of anti-Müllerian hormone (AMH), a glycoprotein hormone that is synthesized exclusively by ovarian granulosa cells, as a sensitive noninvasive biomarker for diagnosing GTCTs in cattle. To achieve this aim, we conducted two experiments. In experiment 1, four clinically healthy Japanese Black cows had blood samples taken daily over one estrous cycle to characterize their AMH profiles throughout the estrous cycle. Additionally, single blood samples were collected from 21 cyclic cows to clarify the physiological range of AMH. In experiment 2, cows with histologically confirmed GTCT (GTCT group, n = 9) and cows affected with cystic ovarian disease (COD group, n = 8) had one blood sample taken before extraction of the tumorous ovary or therapeutic treatment for the COD. Blood samples (n = 105) from cyclic cows (n = 25) in experiment 1 were assigned as a physiologically cyclic group (PC group). Plasma AMH, immunoreactive inhibin (ir-INH), estradiol-17β (E2), testosterone (T), and progesterone (P4) concentrations were assayed in all samples. In experiment 1, the mean plasma AMH concentration was 0.09 ± 0.003 ng/mL and did not show substantial fluctuation throughout the estrous cycle. In experiment 2, plasma AMH, ir-INH, and E2 concentrations were significantly elevated in the GTCT group in comparison with the PC group; among these parameters, only the AMH concentrations were significantly higher in the GTCT group than in the COD group. The area under the receiver operating characteristic curve of AMH for diagnosis of GTCT was 0.99 and was significantly higher than that of ir-INH (P < 0.001) and E2 (P < 0.01). Moreover, the AMH at a cutoff point of ≥0.36 ng/mL had the highest diagnostic accuracy (99.2%), sensitivity (100%), and specificity (99.1%) compared with the other tested parameters. In conclusion, plasma AMH concentration is probably a more reliable and sensitive biomarker for bovine GTCTs than the concentrations of ir-INH or ovarian steroids.

Risk factors for stillbirth and dystocia in Japanese Black cattle.

Stillbirth and dystocia are major factors reducing the productivity of beef cattle. The objective of this study was to determine the effect of season, parity and gestation length on the rates of stillbirth and dystocia in Japanese Black cattle. Calving records were obtained from 905 farms in Miyazaki Prefecture, Japan. Data were collected from 41,116 calvings in 15,378 (14.42% primiparous). There were 1013 stillbirths (2.46%) and 3514 dystocias (8.55%). The stillbirth rate in winter (December to February) (3.18%) was higher (odds ratio (OR) [95% confidence interval]: 1.008 [1.004-1.012]) than that in summer (June to August). Similarly, the dystocia rates in winter (OR: 1.011 [1.004-1.019]) and spring (March to May) (OR: 1.020 [1.013-1.027]) were significantly higher than in summer. For primiparous cows, the rates of stillbirth (OR: 1.010 [1.004-1.015]) and dystocia (OR: 1.053 [1.042-1.064]) were higher than in cows with fifth parity (reference parity). Stillbirth rates were higher in cows at ≥ 301 days of pregnancy (OR: 1.049 [1.035-1.062]) and those at ≤ 270 days of pregnancy (OR: 2.072 [2.044-2.101]) than those at between 281 and 290 days of pregnancy. Likewise, dystocia rates were higher in cows at ≥ 301 days of pregnancy (OR: 1.033 [1.008-1.059]) and those at ≤ 270 days of pregnancy (OR: 1.124 [1.095-1.154]) than those at between 281 and 290 days of pregnancy. Winter, primiparity, and long and short gestation lengths were risk factors for stillbirth and dystocia in this cohort of Japanese Black cattle.

Mycoplasma infection in the uterus of early postpartum dairy cows and its relation to dystocia and endometritis

This study investigated the incidence of mycoplasma infection in the uterus of postpartum Holstein dairy cows and its relationship to the occurrence of endometritis. The genital tracts of 209 cows from three dairy farms in the Iwate Prefecture, Japan, were examined at Weeks 5 and 7 postpartum. The condition of the cervicovaginal mucus was assessed using a Metricheck device and assigned a score from 0 (clear mucus) to 4xa0(purulent material with fetid odor). Intrauterine samples (N = 418) were collected at Weeks 5 and 7 postpartum using a cytobrush. After its withdrawal, swab samples were placed in mycoplasma culture broth at 37 °C for 72 hours. A novel and rapid polymerase chain reaction was used to detect seven mycoplasma species (Mycoplasma bovis, M.xa0arginini, M. bovigenitalium, M. californicum, M. bovirhinis, M. alkalescens, and M. canadense). The cytobrush was also rolled gently along the length of a glass slide for subsequent polymorphonuclear neutrophil count. The diagnostic criteria for cytological endometritis were 6% or more and 4% or more polymorphonuclear neutrophils at Weeks 5 and 7, respectively. From a subset of cows, additional swabs were rolled against the cytobrush and then placed in transport medium. These samples were then plated on specific agar plates and cultured under aerobic and anaerobic conditions to identify other bacteria present. The incidence of dystocia at the last calving was compared in mycoplasma positive and negative cows. Of the seven mycoplasma species, only M. bovigenitalium was detected; it was detected in 31 of the 418 uterine swabs (7.4%). Twenty-four cows were positive for M. bovigenitalium (eight cows at Week 5, nine cows at Week 7, and seven cows at both Weeks 5 and 7). The incidence of dystocia was higher (P < 0.0001) in mycoplasma positive (7/24; 29.2%) compared with mycoplasma negative (4/185; 2.2%) cows. However, there was no significant association between dystocia at last calving and subsequent uterine infection with other bacteria. In addition, the incidence of cytologic endometritis was higher (P < 0.05) in mycoplasma positive (8/16; 50%) than in mycoplasma negative (47/193; 24.4%) cows at Week 7. Therefore, we concluded that M. bovigenitalium infection in the uterus might be associated with recent dystocia and with cytologic endometritis in postpartum dairy cows.

Influences of Metabolic Traits on Subclinical Endometritis at Different Intervals Postpartum in High Milking Cows

Seventy pluriparous high-yielding cows were used to investigate the impact of metabolic traits and body condition score (BCS) during early lactation on subclinical endometritis diagnosed at weeks 5, 6 and 7 postpartum (pp). Blood samples were collected from animals with no peripartum problems from the second (W2) to seventh (W7) weeks pp to estimate some blood metabolites including non-esterified fatty acids (NEFA), β-hydroxybutyric acid (BHBA), blood glucose, total cholesterol (T-chol) and blood urea nitrogen (BUN). Reproductive tract examination was carried out at weeks 5, 6 and 7 pp by endometrial cytology (percentage of polymorphonuclear cells; PMN%). Based on PMN%, animals having <5% were defined as subclinical endometritis group (ENDM group) while animals unaffected by endometritis were defined as no subclinical endometritis group (NOENDM group). Animals with endometritis during week 5 were identified as ENDM5, during week 6 identified as ENDM6 and during week 7 identified as ENDM7 or animals with no endometritis during weeks 5 (NOENDM5), 6 (NOENDM6) and 7 (NOENDM7) pp. Animals diagnosed at week 5; BUN and BCS were lower p < 0.05 in ENDM 5 than NOENDM5 group at W2, W4, W6 and W7. Cows diagnosed at week 6; T-chol was significantly higher (p = 0.05) in ENDM6 group (279.2 ± 12.5 mg/dl) than NOENDM6 group (246 ± 9.5 mg/dl) at W7. Moreover, blood glucose was significantly low (p < 0.05) in ENDM6 group when compared to NOENDM group at W4u2003pp (49.2 ± 1.8 vs 53.8 ± 1.3 mg/dl). BCS was significantly lower (p < 0.001) in animals suffered from endometritis during week 7 when compared to NOENDM7 cows at W3, W4, W5, W6 and W7. In conclusion, lower blood glucose, BUN and BCS could be a risk factor for cytologically diagnosed endometritis at weeks 5, 6 and 7 pp.

Effects of stillbirth and dystocia on subsequent reproductive performance in Japanese Black cattle.

The objective of this study was to determine the effects of stillbirth and dystocia on subsequent reproductive performance in Japanese Black cattle. Data were collected for 34,763 calvings from 13,186 animals on 826 farms. Stillbirth was associated with a decreased first service conception rate compared to cows with a normal calving (44.4 versus 54.0%, respectively, Pu2009<u20090.001). Cows with a normal calving required fewer artificial inseminations per conception and had a lower stillbirth rate at subsequent parity than those with stillbirth and dystocia (Pu2009<u20090.001). Japanese Black cows with stillbirth and dystocia had inferior subsequent reproductive performance compared with those with a normal calving.

Seroepidemiological survey of sheep flocks from Northern Japan for Mycoplasma ovipneumoniae and Mycoplasma agalactiae

Sheep flocks from Hokkaido, Iwate and Aomori, three northern prefectures of Japan, were screened for antibodies to Mycoplasma ovipneumoniae and Mycoplasma agalactiae by ELISA. Sixty four animals out of 246 (26%) were seropositive to M. ovipneumoniae, with positive results obtained from all three prefectures. None of the sera tested were serologically positive to M. agalactiae.

Prevalence of antibodies against Parainfluenza virus type 3, Respiratory syncitial virus and bovine Herpesvirus type 1 in sheep from Northern Prefectures of Japan.

Ovine sera collected in the Prefectures of Hokkaido, Aomori and Iwate in the Northern Japan were examined for the presence of antibodies against Respiratory syncytial virus (RSV), bovine Herpesvirus type 1 (infectious bovine rhinotracheitis: IBR) and Parainfluenza virus type 3 (PIV3) using serum neutralisation (SN) and enzyme-linked immunosorbent assay (ELISA) tests. Twenty-three animals (11.73%) out of the 196 tested were sero-positive to PIV3. Sixteen animals (8.69%) out of the 184 tested reacted to RSV. No animals were positive to IBR antigen. Sero-conversions to PIV3 were detected in Hokkaido and Iwate (14.92% and 8.82%, respectively). Antibodies against RSV were detected in Hokkaido (9.23%) and Aomori (14.28%). Although no diagnostic measures were in place, the infections did not appear to be related to any reduction in sheep productivity.

Reproductive performance of Japanese Black cattle: Association with herd size, season, and parity in commercial cow-calf operations

The Japanese Black is the most common breed of beef cattle in Japan. However, only limited data are available on the associations of season, parity, and herd size with reproductive performance in Japanese Black cattle. Therefore, the objective of the present study was to determine the associations of these factors with reproductive performance parameters, such as the calving to first service interval (CFSI) and first service conception rate in Japanese Black cattle. Data were collected from 34,763 calvings in 13,186 animals from 826 commercial cow-calf operations in the Miyazaki prefecture, which is located on the south eastern coast of Kyushu, Japan. This region has a temperate climate with warm humid summers and cold winters. All cattle were reared intensively, and the animals were housed in free stalls throughout their lives. The mean number of cows per farm was 18 (range, 1-454). All animals were bred by artificial insemination. Herds were classified into three groups based on size: small (≤10 cows), intermediate (11-50 cows), and large (≥51 cows). The mean (±SD) parity, CFSI, and the first service conception rate were 4.9xa0±xa02.9, 80.0xa0±xa046.2xa0days, and 53.5 ±xa049.9%, respectively. Cows that calved in the spring (March to May) and winter (December to February) had the longest CFSI (Pxa0<xa00.05). The CFSI in first-parity cows was shorter than in cows at parity 7 or higher (Pxa0<xa00.05). Cows in large herds had an approximately 10xa0days shorter mean CFSI than those in small herds (Pxa0<xa00.05). Cows inseminated in the winter or spring had an approximately 5% points lesser first-service conception rate (FSCR) than those inseminated during the summer (June to August) or autumn (September to November; Pxa0<xa00.05). As parity increased from 1 to 9, FSCR decreased from 60.0% to 43.1% (Pxa0<xa00.05). Cows in small herds had a lesser FSCR than those in intermediate and large herds (Pxa0<xa00.05). In summary, decreased reproductive performance in intensively reared Japanese Black cattle was associated with calving and artificial insemination during the winter and spring, greater parity, and small herd size.

Serum selenium and liposoluble vitamins in Japanese Black cows that had stillborn calves.

Stillbirth and dystocia are major factors that negatively affect beef production. We sought to clarify serum selenium and liposoluble vitamin levels in Japanese Black cows that gave birth to stillborn calves (stillbirth cows). Blood samples were collected from 103 stillbirth cows and 95 cows that gave birth to healthy calves (control cows). Serum levels of selenium (45.8 ± 16.0 ng/ml) and vitamin A (73.0 ± 24.8 IU/dl) in stillbirth cows were lower (P<0.05) than those in control cows (52.2 ± 8.9 ng/ml and 93.3 ± 14.8 IU/dl, respectively). Our findings suggest that appropriate serum selenium and vitamin A levels are important for calving cows.

Evidence of Spontaneous Recovery of Granulosa-Theca Cell Tumour in a Heifer: A Retrospective Report.

A 14-month-old Japanese Black heifer was evaluated on Day 0 (D 0) for enlargement of the right ovary (RO). Transrectal ultrasonography (TRUS) revealed that the RO was markedly enlarged and multicystic, while the left ovary (LO) was small and inactive. The presumptive diagnosis was granulosa-theca cell tumour (GTCT), which was confirmed by markedly elevated plasma anti-Müllerian hormone (AMH) of 4.42xa0ng/ml. Therefore, ovariectomy of the RO was the treatment of choice. The heifer was checked by TRUS and blood sampling on D 42, D 63 and immediately before ovariectomy on D 85. On D 42, TRUS did not show marked changes in either ovary in comparison with D 0. However, on D 63, the RO had transformed into a single cyst, and on D 85, the LO had resumed cyclic activity. The RO was extracted on D 85 by hand-assisted laparoscopic ovariectomy to allow better control. Unexpectedly, histopathology revealed the lesion to be a fluid-filled cystic structure, with no neoplastic proliferation of follicular epithelium that would indicate GTCT. The wall of the cystic structure consisted of collagen fibres and a few degenerated granulosa cells. The retrospective hormonal analysis revealed that the AMH concentrations had markedly dropped on D 63 and 85, which coincided with resumption of cyclicity in the LO. These findings suggest that the GTCT had self-cured and transformed into a cyst-like structure. The heifer then received an oestrous synchronization regime on D 105, was artificially inseminated on D 115 and became pregnant.