Antimicrobial resistance in Klebsiella species from milk specimens submitted for bovine mastitis testing at the Wisconsin Veterinary Diagnostic Laboratory, 2008-2019.

The purpose of this retrospective study was to describe antimicrobial resistance trends in Klebsiella isolates cultured from milk samples submitted to the Wisconsin Veterinary Diagnostic Laboratory for bovine mastitis testing. In vitro antimicrobial susceptibility testing was conducted in 483 Klebsiella isolates cultured from 63,841 milk samples submitted from January 1, 2008, through December 31, 2019. The Wisconsin Veterinary Diagnostic Laboratory conducted antimicrobial susceptibility testing according to the Clinical and Laboratory Standards Institute by using a broth microdilution panel. Ten antimicrobials were tested in the panel: ampicillin, penicillin, erythromycin, oxacillin + 2% NaCl, pirlimycin, penicillin/novobiocin, tetracycline, ceftiofur, cephalothin, and sulfadimethoxine. Isolates were considered resistant to specific antimicrobials based on minimum inhibitory concentrations described in Clinical and Laboratory Standards Institute guidelines. No guidelines were available for sulfadimethoxine; isolates were categorized as resistant when bacterial growth was not inhibited. The proportion of isolates resistant to ceftiofur, cephalothin, or tetracycline did not increase over time. For sulfadimethoxine, the proportion of resistant isolates decreased over time. These results do not demonstrate a trend toward increasing antimicrobial resistance among Klebsiella isolates. Despite that, antimicrobial resistance should continue to be monitored.

Molecular epidemiology of nonsevere clinical mastitis caused by Klebsiella pneumoniae occurring in cows on 2 Wisconsin dairy farms.

The objective of this study was to describe diversity of Klebsiella pneumoniae isolated from milk collected at detection of nonsevere (abnormal milk or abnormal udder) clinical mastitis (CM) and during a follow-up period. Cases were detected in cows enrolled in a randomized clinical trial (n = 168) conducted using 2 related Wisconsin dairy farms. Cases were randomly assigned to receive 2 d (n = 18) or 8 d (n = 18) of intramammary infusions with an approved product containing ceftiofur hydrochloride or assigned to a negative control group (n = 17). Milk samples were collected from affected quarters at detection and during a 28-d follow-up period. Sufficient DNA was recovered from 53 of 54 Kleb. pneumoniae cultured from quarter milk samples collected at detection of the incident case. Additional Kleb. pneumoniae were recovered from milk samples collected from the same quarters at 14, 21, and 28 d after case detection (n = 35), at detection of recurrent cases in the same quarter (n = 14), and from new cases of CM (n = 3) occurring in enrolled quarters. All Kleb. pneumoniae were used for molecular typing by pulsed-field gel electrophoresis and 90% similarity was used to define homology. Of Kleb. pneumoniae recovered from incident cases, unique strains (n = 41) were identified in milk samples collected from cows on farm A (n = 19) and farm B (n = 22), whereas 12 clonal strains were identified with 8 found only in milk collected from farm A and 4 found in milk samples collected from cows on both farms. Heterogeneous strains of Kleb. pneumoniae genotypes were isolated from incident cases of CM. However, when intramammary infection persisted or when recurrence of CM occurred, clonal strains were isolated at 14, 21, or 28 d. Similar strains of Kleb. pneumoniae genotypes caused persistent CM. In conclusion, initial cases of CM were caused by a wide genetic diversity of Kleb. pneumoniae, but when IMI persisted, the same strain often persisted within the mammary gland up to 28 d.

Short communication: Longitudinal study of quarter-level somatic cell responses after naturally occurring, nonsevere clinical mastitis diagnosed as culture negative, or caused by Escherichia coli or Klebsiella pneumoniae, and randomly assigned to a no-treatment group or to receive intramammary ceftiofur.

The objective of this study was to describe weekly quarter-level somatic cell count (QSCC) after occurrence of nonsevere clinical mastitis (CM) that was diagnosed as culture negative, or caused by Escherichia coli or Klebsiella pneumoniae. All cases occurred in cows enrolled in negatively, controlled randomized clinical trials. We hypothesized that after occurrence of CM, QSCC patterns would vary among etiologies and this effect would not be mitigated by treatment using intramammary (IMM) ceftiofur. Data from two previously published randomized clinical trials performed on 3 Wisconsin dairy farms were used. Only cases confirmed as culture negative (NG) or E. coli or Kleb. pneumoniae (GRAMNEG) were used for analysis. In NG, cows were assigned to no antimicrobial treatment (negative control, n = 44) or 5 d of once daily IMM (n = 41) infusions with an approved product containing ceftiofur hydrochloride. In GRAMNEG, cows were assigned to IMM treatment with the same ceftiofur product for 2 different durations (2 d, n = 36; or 8 d, n = 38) or no antimicrobial treatment (negative control, n = 36). For quarters enrolled in NG, no significant differences were identified for weekly QSCC between quarters in the treated or negative control groups (5.4 log10SCC for both groups). For quarters enrolled in GRAMNEG, no significant differences were identified for QSCC between quarters that received the 2-d (6.2 log10SCC) or 8-d (6.3 log10SCC) IMM treatment or were in the negative control group (6.0 log10SCC). At the pathogen level, regardless of treatment, QSCC varied among pathogens and log10SCC were 5.4 (culture negative), 5.8 (E. coli), and 6.2 (Kleb. pneumoniae). Patterns of QSCC of CM diagnosed as culture negative and E. coli were similar in magnitude and time to resolution of the inflammatory response. In conclusion, as compared to CM diagnosed as culture negative or caused by E. coli, CM caused by Kleb. pneumoniae was associated with poorer outcomes. Regardless of IMM ceftiofur treatment, the immune response of the cow resulted in rapid reduction of SCC of quarters diagnosed as culture negative and quarters with CM caused by E. coli. In contrast, the SCC remained elevated for quarters with CM caused by Kleb. pneumoniae and a greater proportion of those cases remained chronically infected.

Negatively controlled, randomized clinical trial to evaluate intramammary treatment of nonsevere, gram-negative clinical mastitis.

The objective of this negatively controlled, randomized clinical trial was to examine clinical outcomes of 2-d or 8-d treatment using an approved intramammary (IMM) product containing ceftiofur hydrochloride compared with no antimicrobial treatment of nonsevere, gram-negative cases of clinical mastitis (CM). Additionally, we contrasted clinical outcomes of cases caused by Escherichia coli (n = 56) or Klebsiella pneumoniae (n = 54). Cases (n = 168) of nonsevere (abnormal milk or abnormal milk and udder) CM were randomly assigned to receive 2 d (n = 56) or 8 d (n = 56) of IMM ceftiofur or assigned to a negative control group (n = 56). At enrollment, quarter milk samples were collected and used for on-farm culture, somatic cell count (SCC), and confirmatory microbiological analysis. Quarter milk samples were collected weekly from 7 to 28 d after enrollment for microbiological and SCC analysis. Clinical outcomes were followed for 90 d or until the end of lactation (follow-up period, FUP). Overall, no significant differences in quarter-level recurrence of CM (32% for negative control, 34% for the 2-d treatment, and 32% for the 8-d treatment), culling (18% for negative control, 12% for 2-d treatment, and 11% for 8-d treatment), voluntary dry-off of affected quarters (20% for negative control, 30% for 2-d treatment, and 27% for 8-d treatment), days until return to normal milk (4.2 days for negative control, 4.8 days for 2-d treatment, 4.5 days for 8-d treatment), weekly quarter-SCC during the FUP (6.1, 6.3, and 6.0 log10SCC for the negative control, 2-d, and 8-d treatments, respectively), or daily milk yield during the FUP (37.1, 36.3, and 37.6 kg/cow per day for the negative control, 2-d, and 8-d treatments, respectively) were observed among experimental groups. Days of discarded milk were greater for cows assigned to 8-d IMM ceftiofur (11.1 d) than for cows assigned to 2-d (6.9 d) or cows assigned to negative control (5.6 d). Bacteriological cure (BC) at 14 and 21 d after enrollment was greater in cows assigned to 8-d (89%) and 2-d (84%) treatment than in cows assigned to negative control (67%), but this outcome was confounded by pathogen. For CM caused by Kleb. pneumoniae, BC was greater for quarters assigned to receive treatment (combined 2-d and 8-d groups; 74% BC) than for quarters assigned to negative control (18%). In contrast, no differences in BC were observed for CM caused by E. coli (97-98%). Culling and voluntary dry-off of affected quarters were significantly greater for cows with quarters affected by Kleb. pneumoniae (22% culled, 39% voluntary dry-off of quarters) than for cows with quarters affected with E. coli (7% culled, 11% voluntary dry-off of quarters). Overall, use of IMM ceftiofur did not result in improvement of most clinical outcomes, but differences between E. coli and Kleb. pneumoniae were evident. In contrast to E. coli, Kleb. pneumoniae caused chronic intramammary infection and induced worse clinical outcomes. Intramammary antibiotic treatment of most mild and moderate cases of CM caused by E. coli is not necessary, but more research is needed to identify which quarters affected by Kleb. pneumoniae may benefit from antimicrobial therapy.

Negatively controlled, randomized clinical trial to evaluate use of intramammary ceftiofur for treatment of nonsevere culture-negative clinical mastitis.

The objective of this negatively controlled randomized clinical trial was to compare clinical outcomes of 5-d intramammary treatment using ceftiofur hydrochloride and no antimicrobial treatment of nonsevere culture-negative cases of clinical mastitis (CM). A total of 121 cases of nonsevere (abnormal milk or abnormal milk and udder) culture-negative CM were randomly assigned to either treatment (n = 62) or negative control (n = 59) groups. Quarters assigned to treatment received 1 daily intramammary infusion with an approved commercially available product containing ceftiofur hydrochloride for 5 d. Quarters assigned to the negative control group did not receive any interventions. Enrolled cows were followed for 90 d or until the end of lactation. At enrollment, milk samples from the affected quarter were used for on-farm culture, somatic cell count (SCC) analysis, and further microbiological analysis. During the follow-up period, milk samples were collected for microbiological analysis and SCC analysis. No significant differences between treatment and negative control groups were identified for treatment failure (5% for treatment vs. 10% for negative control, n = 121), quarter-level CM recurrence (8 vs. 5%, n = 91), intramammary infection at 14 or 28 d after enrollment (13 vs. 26%, n = 86), days until clinical cure (4.2 vs. 4.0 d), days to culling (48.3 vs. 36.8 d), daily milk production (43.3 vs. 43.6 kg/cow per day), or weekly quarter SCC (5.5 vs. 5.4 log10 SCC). Days of milk discard were greater for cows assigned to the treatment group (8.5 d) compared with cows assigned to the negative control group (5.6 d). During the follow-up period, cases in the treatment group had a 50% risk reduction in IMI compared with cases in the negative control group. Irrespective of group, negative outcomes such as quarter-level CM recurrence (12%), treatment failure (12%), and culling (5%) occurred infrequently in nonsevere culture-negative cases of CM. Use of intramammary ceftiofur for treatment of nonsevere culture-negative cases of CM did not improve any economically relevant clinical outcome such as culling, milk production, or SCC.

Modifications to Ovsynch improve fertility during resynchronization: Evaluation of presynchronization with gonadotropin-releasing hormone 6 d before initiation of Ovsynch and addition of a second prostaglandin F2α treatment.

Lactating Holstein cows (n=897) were randomly assigned to a 2×2 factorial arrangement of treatments to compare the main effects of presynchronization with GnRH 6 d before beginning an Ovsynch protocol (±GnRH) and a second PGF2α treatment 24h after the first (1 vs. 2 PGF2α) on pregnancies per artificial insemination (P/AI). This resulted in the following 4 treatments: (1) an Ovsynch protocol (GPG, control); (2) presynchronization with GnRH followed by an Ovsynch protocol (GGPG); (3) an Ovsynch protocol with a second PGF2α treatment (GPPG); and (4) presynchronization with GnRH followed by an Ovsynch protocol with a second PGF2α treatment (GGPPG). All cows were submitted for first timed artificial insemination (TAI) using a Presynch Ovsynch protocol, and cows detected in estrus after the second PGF2α treatment of the Presynch portion of the protocol were inseminated and removed from the experiment. Nonpregnant cows were resynchronized using an Ovsynch protocol initiated 32±3 d after artificial insemination. Blood samples were collected at the first GnRH treatment (G1), at the PGF2α treatment (PGF), and at the last GnRH treatment (G2) of the Ovsynch protocol and were assayed for progesterone (P4) concentrations. Overall, P/AI tended to be greater for cows receiving a second PGF2α treatment compared with cows not receiving the second PGF2α treatment (40 and 37% for GGPPG and GPPG treatments, respectively, vs. 33 and 32% for GGPG and GPG treatments, respectively). Interestingly, treatment effects on P/AI were detected only for resynchronized cows receiving second and greater TAI, but not for cows receiving first TAI. Fewer cows presynchronized with GnRH had low (<0.5ng/mL) P4 at G1 than cows not presynchronized (13 vs. 25%), and P4 was greater at PGF2α for cows presynchronized with GnRH (4.4 vs. 4.0ng/mL). Cows receiving 2 PGF2α treatments had lower P4 at G2 than cows receiving 1 PGF2α treatment (0.15 vs. 0.35ng/mL). Differences in P4 at PGF2α were detected only for resynchronized cows and not for cows submitted for first TAI. We conclude that presynchronization with GnRH 6 d before beginning an Ovsynch protocol tended to increase P4 at the first GnRH treatment of an Ovsynch protocol, and that a second PGF2α treatment 24h after the first decreased P4 at TAI, thereby increasing P/AI in cows resynchronized for second and greater TAI.

The association between occurrence and severity of subclinical and clinical mastitis on pregnancies per artificial insemination at first service of Holstein cows.

The objective of this prospective study was to determine associations between occurrence and severity of clinical (CM) and subclinical mastitis (SM) during a defined breeding risk period (BRP, 3d before to 32d after artificial insemination) on pregnancies per artificial insemination at first service (P/AI1). Dairy cows (n=3,144) from 4 Wisconsin herds were categorized based on the occurrence of one or more CM or SM events during and before the BRP: (1) healthy, (2) mastitis before BRP, (3) SM during BRP, (4) chronic SM, (5) CM during BRP, or (6) chronic CM. Clinical mastitis cases were categorized based on etiology (gram-negative, gram-positive, and no growth) and severity (mild, moderate, or severe). Compared with healthy cows, the odds of pregnancy were 0.56, 0.67, and 0.75 for cows experiencing chronic CM, CM, or SM during the BRP, respectively. The occurrence of chronic SM was not associated with reduced probability of P/AI1. Compared with healthy cows, the odds of pregnancy were 0.71 and 0.54 for cows experiencing mild or moderate-severe cases of CM during the BRP, respectively. The odds of pregnancy for cows experiencing CM caused by gram-negative or gram-positive bacteria during the BRP were 0.47 and 0.59, respectively. The occurrence of CM that resulted in no growth of bacteria in cultured milk samples was not associated with reductions in P/AI1. Regardless of etiology, microbiologically positive cases of CM with moderate or severe symptoms were associated with substantial reductions in P/AI1. Etiology, severity, and timing of CM were associated with decreases in the probability of pregnancy at first artificial insemination. Severity of the case was more important than etiology; however, regardless of severity, microbiologically negative cases were not associated with reduced probability of pregnancy.

Presynchronization using a modified Ovsynch protocol or a single gonadotropin-releasing hormone injection 7 d before an Ovsynch-56 protocol for submission of lactating dairy cows to first timed artificial insemination.

Presynchronization strategies, such as Presynch-Ovsynch and Double-Ovsynch, increase fertility to timed artificial insemination (TAI) compared with Ovsynch alone; however, simpler presynchronization strategies could reduce costs and simplify reproductive management. Lactating Holstein cows (n=601) were randomly assigned to 1 of 2 presynchronization treatments before beginning an Ovsynch-56 protocol (GnRH at 70 ± 3 DIM, PGF2α 7 d later, GnRH 56 h after PGF2α, and TAI 16 h later at 80 ± 3 DIM) for first TAI. Cows (n=306) in the first treatment (Double-Ovsynch; DO) were presynchronized using a modified Ovsynch protocol (GnRH at 53 ± 3 DIM, 7 d later PGF2α, and GnRH 3 d later) ending 7 d before the first GnRH injection (G1) of an Ovsynch-56 protocol. Cows (n=295) in the second treatment (GGPG) were presynchronized using a single injection of GnRH 7 d before G1 of an Ovsynch-56 protocol at 63 ± 3 DIM. Blood samples were collected at G1 and the PGF2α injections of the Ovsynch-56 protocol to determine progesterone (P4) concentrations. Pregnancy diagnosis was performed using ultrasonography 32 d after TAI, and pregnant cows were reexamined 46 and 70 d after TAI. Overall, DO cows had more pregnancies per artificial insemination (P/AI) compared with GGPG cows 32 d after TAI (53 vs. 43%). Overall, P/AI did not differ by parity (primiparous vs. multiparous), and pregnancy loss did not differ between treatments or parities. More DO cows had P4 in a medium range (>0.5 to <4 ng/mL) at G1 of the Ovsynch-56 protocol compared with GGPG cows (82 vs. 50%), and more DO cows had high P4 (>4 ng/mL) at the PGF2α injection of the Ovsynch-56 protocol compared with GGPG cows (67 vs. 36%). Thus, presynchronization with a modified Ovsynch protocol increased P/AI after TAI at first AI by increasing synchrony to the Ovsynch-56 protocol compared with presynchronization using a single injection of GnRH.

Relationships between fertility and postpartum changes in body condition and body weight in lactating dairy cows.

The relationship between energy status and fertility in dairy cattle was retrospectively analyzed by comparing fertility with body condition score (BCS) near artificial insemination (AI; experiment 1), early postpartum changes in BCS (experiment 2), and postpartum changes in body weight (BW; experiment 3). To reduce the effect of cyclicity status, all cows were synchronized with Double-Ovsynch protocol before timed AI. In experiment 1, BCS of lactating dairy cows (n = 1,103) was evaluated near AI. Most cows (93%) were cycling at initiation of the breeding Ovsynch protocol (first GnRH injection). A lower percentage pregnant to AI (P/AI) was found in cows with lower (≤ 2.50) versus higher (≥ 2.75) BCS (40.4 vs. 49.2%). In experiment 2, lactating dairy cows on 2 commercial dairies (n = 1,887) were divided by BCS change from calving until the third week postpartum. Overall, P/AI at 70-d pregnancy diagnosis differed dramatically by BCS change and was least for cows that lost BCS, intermediate for cows that maintained BCS, and greatest for cows that gained BCS [22.8% (180/789), 36.0% (243/675), and 78.3% (331/423), respectively]. Surprisingly, a difference existed between farms with BCS change dramatically affecting P/AI on one farm and no effect on the other farm. In experiment 3, lactating dairy cows (n = 71) had BW measured weekly from the first to ninth week postpartum and then had superovulation induced using a modified Double-Ovsynch protocol. Cows were divided into quartiles (Q) by percentage of BW change (Q1 = least change; Q4 = most change) from calving until the third week postpartum. No effect was detected of quartile on number of ovulations, total embryos collected, or percentage of oocytes that were fertilized; however, the percentage of fertilized oocytes that were transferable embryos was greater for cows in Q1, Q2, and Q3 than Q4 (83.8, 75.2, 82.6, and 53.2%, respectively). In addition, percentage of degenerated embryos was least for cows in Q1, Q2, and Q3 and greatest for Q4 (9.6, 14.5, 12.6, and 35.2% respectively). In conclusion, for cows synchronized with a Double-Ovsynch protocol, an effect of low BCS (≤ 2.50) near AI on fertility was detected, but change in BCS during the first 3 wk postpartum had a more profound effect on P/AI to first timed AI. This effect could be partially explained by the reduction in embryo quality and increase in degenerate embryos byd 7 after AI in cows that lost more BW from the first to third week postpartum.

The transition between preluteolysis and luteolysis in cattle.

Novel characterization of the transition between preluteolysis and luteolysis was done in seven heifers. Blood samples were collected hourly and assayed for progesterone (P4), 13-14-dihydro-15-keto-PGF2α (PGFM), and estradiol (E2). The peaks of P4 oscillations were used to designate the transitional hour in each heifer. The interval from the peak of the last PGFM pulse of preluteolysis to the peak of the first pulse during luteolysis (transitional period) was longer (P < 0.0001) than the interval between the first and second pulses during luteolysis (13.4 ± 1.3 h vs. 7.0 ± 0.9 h). The long intervals from the last PGFM pulse of preluteolysis to the transitional hour (4.0 ± 0.9 h) and from the transitional hour to the first PGFM pulse of luteolysis (9.4 ± 1.3 h) resulted in the illusion that the beginning of luteolysis was not associated temporally with a PGFM pulse. The E2 and PGFM concentrations were less (P < 0.05) during the last PGFM pulse of preluteolysis than during the first pulse of luteolysis. Concentration of P4 was suppressed at the peak of the last PGFM pulse of preluteolysis and consistently rebounded at the transitional hour to the concentrations before the PGFM pulse. In four of seven heifers, one or two P4 rebounds occurred between the peak of the PGFM pulse and the rebound at the transitional hour. Results indicated that the prolonged transitional period may be related, at least in part, to increasing concentration of E2, intervening P4 rebounds between the peak of the last PGFM pulse of preluteolysis and the transitional hour, and the complete P4 rebound at the transitional hour.

Effect of luteinizing hormone oscillations on progesterone concentrations based on treatment with a gonadotropin-releasing hormone antagonist in heifers.

Close temporality has been reported between the episodic secretion of luteinizing hormone (LH) and progesterone (P4) during the midluteal phase and preceding the beginning of luteolysis in cattle. In the present studies, the relationship between LH and P4 was examined by blocking LH oscillations with the gonadotropin-releasing hormone (GnRH) antagonist, acyline. In a titration study, the minimal single acyline dose for blocking LH oscillations in heifers was 3 µg/kg. The main experiment compared LH and P4 concentrations and oscillations between a group treated with acyline on day 15 after ovulation (n = 8) and a control group (n = 4). Concentrations of P4 in blood samples collected every 8 h on days 13 to 18 indicated that acyline treatment did not alter the time that luteolysis began or the length of the luteolytic process. In blood samples collected every hour for 24 h beginning at the hour of treatment, acyline reduced the LH concentrations and blocked LH oscillations. The hourly LH means were 0.06 to 0.08 ng/mL, comparable to the mean concentration at the nadirs of LH oscillations in controls (0.07 ng/mL). During the hourly sampling, the GnRH antagonist produced the following P4 responses: (1) lower P4 concentrations, (2) fewer and reduced prominence of P4 oscillations, and (3) increased length and variability in the interval between the peaks of P4 oscillations. Results indicated that LH oscillations affect both the prominence and the rhythmicity of P4 oscillations during preluteolysis but not the onset and length of luteolysis.

Concomitance of luteinizing hormone and progesterone oscillations during the transition from preluteolysis to luteolysis in cattle.

The temporal relationships of episodes of luteinizing hormone (LH) oscillations, 13,14-dihydro-15-keto-PGF2α (PGFM) pulses, and progesterone (P4) fluctuations during the latter portion of preluteolysis and the early portion of luteolysis were characterized. In Experiment 1, the detection of LH episodes in blood samples collected every 15 min for 8 h was compared with detection in the samples collected every hour in 4 heifers. The number of independently detected episodes/heifer (total = 7) was the same for the 15-min and hourly collection intervals. In Experiment 2, blood samples were collected every hour (n = 7 heifers) and retrospectively assigned to 15 h before and 15 h after the transitional hour between preluteolysis and luteolysis. During preluteolysis, compared with luteolysis, the amplitude of LH oscillations was greater (0.28 ± 0.03 vs 0.18 ± 0.03 ng/mL; P < 0.02) and the interval between peaks of LH oscillations was shorter (3.3 ± 0.3 h vs 4.3 ± 0.6 h; P < 0.04). The LH peaks occurred at the same hour as the peak of a P4 fluctuation in 77% and 29% of LH oscillations (P < 0.0009) during preluteolysis and luteolysis, respectively. In preluteolysis, synchrony between LH and P4 episodes occurred consistently during the P4 rebound after the peak of a PGFM pulse. In luteolysis, the LH peak preceded the peak of the P4 rebound. On a temporal basis, the hypothesis was supported that episodic LH accounts, at least in part, for the reported P4 rebound that occurs after the P4 suppression at the peak of a PGFM pulse.

Diurnal variation in LH and temporal relationships between oscillations in LH and progesterone during the luteal phase in heifers.

Diurnal variation in progesterone and LH during the luteal phase and the temporal relationships between oscillations of the two hormones were studied in 10 heifers by collection of blood samples at 0100, 0700, 1300, and 1900 h each day, beginning on Day 1 (Day 0 = ovulation). Concentration of LH on Days 5-9, but not on Days 10-14, was lower (P < 0.05) at 0700 h (0.25 ± 0.02 ng/mL) than at each of the other three hours (combined, 0.32 ± 0.02 ng/mL). An oscillation was defined as an uninterrupted increase and decrease in concentrations. The number of LH oscillations/heifer with the peak at 1900 h (6.1 ± 0.7) throughout the luteal phase was greater (P < 0.01) than for each of the other hours (combined, 4.0 ± 0.2). Diurnal variation in progesterone was not detected. Only statistically defined LH oscillations were used to determine the temporal association between the peak of an LH oscillation and various components of a progesterone oscillation. On Days 5-14, the frequency of the peak of an LH oscillation occurring at the same hour as the peak of a progesterone oscillation (26/48, 54%) was greater (P < 0.0001) than at the progesterone nadir (3/48, 6%). The frequency of the LH peak occurring during increasing (11/34, 32%) and decreasing (8/25, 32%) progesterone concentrations was intermediate (P < 0.05). Results indicated the following: 1) diurnal variation occurred in LH as determined by concentration and by the hour of the peak of an oscillation; and 2) LH oscillations were temporally and positively related to progesterone oscillations.

Intrapulse temporality between pulses of a metabolite of prostaglandin F 2α and circulating concentrations of progesterone before, during, and after spontaneous luteolysis in heifers.

Pulses of the prostaglandin F(2α) (PGF) metabolite 13,14-dihydro-15-keto-PGF(2α) (PGFM) and the intrapulse concentrations of progesterone were characterized hourly during the preluteolytic, luteolytic, and postluteolytic periods in seven heifers. The common hour of the end of preluteolysis and the beginning of luteolysis was based on a progressive progesterone decrease when assessed only at the peaks of successive oscillations. The end of the luteolytic period was defined as a decrease in progesterone to 1 ng/mL. Blood samples were taken hourly from 15 d after ovulation until luteal regression as determined by color-Doppler ultrasonography. Between Hours -2 and 2 (Hour 0 = PGFM peak) of the last PGFM pulse of the preluteolytic period, progesterone decreased between Hours -1 and 0, and then returned to the prepulse concentration. Concentration did not change significantly thereafter until a PGFM pulse early in the luteolytic period; progesterone decreased by Hour 0 and transiently rebounded after Hour 0, but not to the prepulse concentration. In the later portion of the luteolytic period, progesterone also decreased between Hours -1 and 0 but did not rebound. After the defined end of luteolysis, progesterone decreased slightly throughout a PGFM pulse. Results demonstrated for the first time that the patterns of progesterone concentrations within a PGFM pulse differ considerably among the preluteolytic, luteolytic, and postluteolytic periods.

Stimulation of pulses of 13,14-dihydro-15-keto-PGF2alpha (PGFM) with estradiol-17beta and changes in circulating progesterone concentrations within a PGFM pulse in heifers.

A single physiologic dose (0.1 mg) of estradiol-17beta in sesame-oil vehicle or vehicle alone (n = 8) was given to heifers on day 14 after ovulation to study the effect on circulating 13-14-dihydro-15-keto-PGF2alpha (PGFM), PGFM pulses, and changes in progesterone concentrations within a PGFM pulse. Blood samples were collected hourly for 16 h after treatment. The estradiol group had a greater mean concentration of PGFM, greater number of heifers with PGFM pulses and number of pulses/heifer, and greater prominence of the PGFM pulses. Changes in progesterone concentrations were not detected during the 16 h sampling session in the vehicle group, indicating that the heifers were in preluteolysis. Progesterone decreased after 12 h in the estradiol group, indicating a luteolytic effect of the estradiol-induced PGF secretion as represented by PGFM concentrations. Intrapulse changes in progesterone were detected during a PGFM pulse in the estradiol group (P < 0.006), but not in the vehicle group. Progesterone increased (P < 0.01) between Hours -2 and -1 of an estradiol-induced PGFM pulse (Hour 0 = peak of pulse), decreased (P < 0.004) between Hours -1 and 0, and increased (P < 0.01) or rebounded between Hours 0 and 1. Results were compatible with previous reports of a role for estradiol in the induction of PGFM pulses in cattle and demonstrated intrapulse changes in progesterone concentrations during an induced PGFM pulse.

Characteristics of pulses of 13,14-dihydro-15-keto-prostaglandin f2alpha before, during, and after spontaneous luteolysis and temporal intrapulse relationships with progesterone concentrations in cattle.

Pulses of the prostaglandin F2alpha (PGF) metabolite 13,14-dihydro-15-keto-PGF (PGFM) were compared among heifers that were in the preluteolytic, luteolytic, and postluteolytic periods (n = 7 or 8 heifers/period). Hourly blood sampling was done in 18-h sessions 15, 16, or 17 days after ovulation. Hourly sampling and statistical identification of a PGFM pulse allowed novel comparisons of PGFM pulses among the three periods. Each period had a similar number of PGFM pulses (2.3 +/- 0.2). The pulses were more prominent during the luteolytic period than during the other periods, as indicated by significantly greater concentration for the peak and amplitude between nadir and peak. Significantly more fluctuations that did not meet the definition of a pulse occurred at the beginning of the preluteolytic period and end of the postluteolytic period than during the luteolytic period. The same nadir ended a pulse and began the next pulse in 85% of adjacent pulses. Seven heifers were selected objectively, based on a progesterone concentration >5 ng/ml at Hour -3 (Hour 0 = peak of PGFM pulse) and a progressive decrease in progesterone from Hours -3 to 0. Progesterone increased (P < 0.03) between Hours 0 and 1, remained at a mean plateau at Hours 1 and 2, and then decreased. Results support the hypothesis of a transient intrapulse rebound in progesterone during an individual PGFM pulse, but only during the first portion of luteolysis. These findings should be considered in future proposals on the mechanisms involved in the effects of PGF on progesterone concentrations.