Effect of treatment of pneumonia and otitis media with tildipirosin or florfenicol + flunixin meglumine on health and upper respiratory tract microbiota of preweaned Holstein dairy heifers.

The objective of this randomized clinical study was to compare the effect of 2 antimicrobial interventions, tildipirosin or florfenicol + flunixin meglumine, used for treatment of pneumonia and extralabel treatment for otitis on health parameters and upper respiratory tract (URT) microbiota of preweaned Holstein calves. Housed preweaned Holstein heifers diagnosed with either otitis or pneumonia were assigned into 1 of 2 treatment groups, receiving a single subcutaneous injection of either 4 mg/kg of tildipirosin (TLD; n = 444) or 40 mg/kg of florfenicol combined with 2.2 mg/kg of a nonsteroidal anti-inflammatory, flunixin meglumine (FLF; n = 442). Calves were enrolled and treated on the day of diagnosis of the first case of pneumonia or otitis. If a calf had a recurrent case, the opposite drug was administered, respecting an interval of 5 d between drug injections. Blood samples for leukocyte counts were collected at 0, 2, 4, and 6 d after treatment, and rectal temperature was measured daily during the 5 d after treatment. Ear scores were observed from calves with otitis. Additionally, swabs of the URT were collected from a subset of 20 calves in each treatment group at d 0, 3, 6, 9, and 11 following enrollment for analysis of URT microbiota through next-generation sequencing of the 16S rRNA gene and quantitative PCR. Swabs were also collected from a comparative group of 20 healthy calves that did not receive any drug. No differences were observed between groups for recurrence risk of either pneumonia (TLD = 32.4%; FLF = 29.7%) or otitis (TLD = 72.7%; FLF = 73.6%). Similarly, no differences were observed for the total number of treatments for pneumonia (TLD = 1.45; FLF = 1.42) or otitis (TLD = 2.96; FLF = 3.07). On the other hand, both drugs reduced rectal temperature, ear scores, and leukocyte counts, with FLF calves having a greater reduction in rectal temperature within 4 d after treatment. Both TLD and FLF reduced the total bacterial load when compared with healthy untreated calves, but no differences were observed between treatment groups. Furthermore, compared with the untreated group, treated calves had lower mean relative abundances (MRA) of the genera Mannheimia, Moraxella, and Pasteurella within 11, 9, and 3 d after treatment, respectively; however, no significant differences were observed between TLD and FLF. On the other hand, MRA of Mycoplasma was not decreased by both treatments compared to untreated animals, and a higher MRA was observed in the TLD group during 11 d after treatment in comparison to FLF and untreated calves. Based on this data, we concluded that both drugs used in the study were effective in reducing rectal temperature, ear scores, leukocyte counts, and MRA of the genera Mannheimia, Pasteurella, and Moraxella in the URT, and calves treated with FLF had a greater reduction in rectal temperature.

Effect of metaphylactic administration of tildipirosin on the incidence of pneumonia and otitis and on the upper respiratory tract and fecal microbiome of preweaning Holstein calves.

The objectives of this study were to evaluate the effect of the metaphylactic use of a semi-synthetic long-acting macrolide (tildipirosin) on the prevention of pneumonia and otitis in preweaning Holstein calves, as well as its effects on the microbiome of their upper respiratory tract (URT) and feces. Newborn healthy Holstein heifers, collectively housed, were randomly allocated to 1 of 2 treatment groups: treatment (TRT; n = 932) or control (CTR; n = 927). Calves in the TRT group received a single subcutaneous injection of 4 mg/kg tildipirosin (Zuprevo, Merck Animal Health) at 7 ± 7 d of life. Calves in the CTR group received no drug injection. All enrolled calves were evaluated from 1 to 63 ± 3 d of life (weaning age) and monitored daily for any adverse health events during this period. Daily physical examination was performed to diagnose pneumonia and otitis, and body weight was measured weekly in all animals. From a randomly selected subset of 217 calves, blood samples for biochemical variables analysis and swabs were collected weekly from the URT and rectum for analysis of the nasal and fecal microbiome, respectively, via next-generation sequencing of the 16S rRNA gene. Total bacterial load was evaluated using quantitative PCR. In addition, another subset of 26 calves was randomly selected and fecal swabs were collected in a more intensive sampling to investigate the short-term effect of tildipirosin administration on the fecal microbiome. We performed general mixed linear models and logistic regression to analyze continuous and binary outcomes, respectively. Tildipirosin metaphylaxis reduced the incidence of otitis (CTR = 47.03%; TRT = 37.55%) and tended to reduce the incidence of pneumonia (CTR = 20.71%; TRT = 17.38%) and the overall mortality risk (CTR = 6.69%; TRT = 4.94%). We observed no significant differences between groups for mortality due to pneumonia (CTR = 0.86%; TRT = 0.97%) or mortality due to otitis (CTR = 2.05%; TRT = 1.39%). Calves in the TRT group had a higher average daily gain than calves in the CTR group. Furthermore, metaphylaxis had no significant effects on the total bacterial load, genus, or phylum analysis of the fecal microbiome from the 2 subset groups. However, for the URT microbiota, we observed a significant decrease in total bacterial load for the TRT group compared to the CTR group 1 week after metaphylactic injection. Tildipirosin metaphylaxis decreased the mean relative abundance of the genera Mannheimia, Moraxella, and Pasteurella but significantly increased the mean relative abundance of Mycoplasma. Although tildipirosin had no positive effect on Mycoplasma, it reduced the mean relative abundance of important pathogenic bacteria in the URT and had positive effects for the control of otitis. The metaphylactic use of tildipirosin can be a suitable strategy for the control of otitis on farms with a high prevalence of this disease.

The Effect of Metaphylactic Use of Tildipirosin for the Control of Respiratory Disease in Long-Distance Transported Dairy Calves.

The objective of this study was to evaluate the efficacy of two metaphylactic strategies using tildipirosin for the control of bovine respiratory disease (BRD) in dairy calves transported to a heifer raising facility within their first week of life. A total of 2,100 calves were enrolled in the study. Animals were transported for ~1,715 km, from dairies located in Minnesota to a calf raising facility located in New Mexico, where they were housed in individual hutches until weaning. Three days after arrival, calves were randomly allocated into three groups: (1) META1: single subcutaneous (SQ) injection of tildipirosin (Zuprevo™, Merck Animal Health) at enrollment at 4 mg/kg; (2) META2: SQ injection of tildipirosin at enrollment and 17 days later; (3) CON: untreated controls. The BRD incidence was 11.4, 10.8, and 9.4% for calves enrolled in the CON, META1, and META2, respectively (P = 0.44). Lung lesions diagnosed through ultrasonography was found in 21.0, 21.0, and 21.8% of calves enrolled in CON, META1, and META2, respectively (P = 0.99). Mortality tended to be greater for CON calves in comparison to META2 calves (1.5 vs. 0.6%, P = 0.06), but did not differ between calves enrolled in CON and META1 groups (1.5 vs. 1.2%, P = 0.55). Growth was not affected by metaphylaxis. The average daily gain for calves enrolled in CON, META1, and META2 was 517, 518 and 525 g, respectively (P = 0.25). Blood analysis revealed that some of the markers of inflammation assessed were influenced by metaphylaxis. At 27 days after enrollment, META2 calves had decreased concentrations of haptoglobin, serum amyloid A, and aspartate aminotransferase, compared to CON calves (P < 0.05). Additionally, CON calves had increased concentrations of globulins and lower albumin to globulin ratio than META2 calves at the end of the weaning period (P < 0.05). In conclusion, tildipirosin metaphylaxis did not decrease the incidence of BRD nor did it have an impact on weight gain. However, metaphylaxis with two injections of tildipirosin at enrollment and 17 days later tended to reduce mortality and improved the systemic inflammatory status of calves during the preweaning period.

Genomic merit for reproductive traits. I: Estrous characteristics and fertility in Holstein heifers.

Genetic selection of dairy cattle in the United States has included reproductive traits (daughter pregnancy rate, DPR; heifer conception rate, HCR), which is believed to have partly contributed to halting the decline in reproductive performance. The objectives of the current study were to evaluate the association among genomic merit for DPR (GDPR) and HCR (GHCR) with estrous characteristics measured by an automated device. Holstein heifers (n = 1,005) were genotyped at 2 mo of age and were classified into quartiles (Q1 = lowest, Q4 = highest) according to the GDPR and GHCR values of the study population. At 10 to 11 mo of age, heifers were fitted with a collar that recorded activity and rumination and determined the occurrence of estrus according to changes in activity and rumination compared with the individual's baseline values. Estrous characteristics of spontaneous estruses (SPE) and PGF2α-synchronized estruses (PGSE) were recorded. Heifers had their estrous cycle synchronized with PGF2α and following detection of estrus received either artificial insemination or embryo transfer according to the herd's genetic selection program. Heifers in Q2 (17.7 ± 0.3 h) of GHCR tended to have longer SPE than heifers in Q4 (16.7 ± 0.3 h). The interaction between GDPR and GHCR was associated with the likelihood of activity peak (0 = no estrus, 100 = maximum activity) ≥80 at SPE because, among heifers in Q3 and Q4 of GHCR, those in Q1 of GDPR were less likely to have an activity peak ≥80. Heifers in Q1 and Q2 of GDPR had reduced hazard of estrus within 7 d of the first PGF2α treatment compared with heifers in Q4 of GDPR. Heifers in Q1 (16.1 ± 0.4 h) of GDPR had shorter PGSE than heifers in Q2 (17.6 ± 0.4 h) and Q4 (17.4 ± 0.4 h) and tended to have shorter PGSE than heifers in Q3 (17.4 ± 0.4 h). Rumination nadir on the day of PGSE was greater for heifers in Q1 (-30.1 ± 0.9 min/d) of GDPR compared with heifers in Q4 (-33.7 ± 0.9 min/d). Among heifers receiving only artificial insemination, those in Q1 of GHCR (adjusted hazard ratio = 0.65; 95% confidence interval = 0.48-0.88) became pregnant at a slower rate than heifers in Q4. Genomic merit for HCR was negatively associated with SPE but tended to be positively associated with hazard of pregnancy, whereas GDPR was positively associated with PGSE and hazard of estrus. Selection of dairy cattle for DPR and HCR may improve reproductive performance through different pathways, namely estrous characteristics and pregnancy establishment.

Estrous characteristics and reproductive outcomes of Holstein heifers treated with 2 prostaglandin formulations and detected in estrus by an automated estrous detection or mounting device.

Dinoprost tromethamine (DIN), a molecule similar to endogenous PGF2α, has a half-life of approximately 9 min. Cloprostenol sodium (CLO), a synthetic analog of PGF2α, has a half-life of approximately 3 h. We hypothesized that treatment of Holstein heifers with CLO would improve estrous detection rate, estrous characteristics, service rate, and overall reproductive performance compared with DIN. Currently in the United States, heifers are largely inseminated based on signs of estrus, which is detected visually or with the aid of mounting detection devices (MD). Automated estrous detection devices (AED) are becoming more accessible to producers, but it is not clear whether they present advantages in the reproductive management of heifers. Therefore, we hypothesized that the use of an AED would improve service and pregnancy rates compared with detection of estrus with the aid of a MD. Holstein heifers (n = 1,019) were enrolled in the experiment at 10 to 11 mo of age, when they were fitted with a Heatime HR LD System (SCR Ltd., Netanya, Israel). At 12 mo of age, we paired heifers according to estrous cycle phase and randomly assigned them to treatments in a 2 × 2 design: PGF2α formulation (CLO vs. DIN) and estrous detection treatment (AED vs. MD). Heifers in the AED treatment were detected in estrus only by the Heatime HR LD System, whereas heifers in the MD treatment were detected in estrus only by the Kamar Heatmount Detector (Kamar Products Inc., Zionsville, IN). Treatments with the same PGF2α formulations were repeated 14 d after the first treatment if heifers had not been detected in estrus. A sub-group of heifers had blood sampled on the day of PGF2α treatment and within 24 h of onset of estrus to determine progesterone and estradiol concentrations. Treatment with CLO reduced the progesterone concentration within 24 h of onset of estrus compared with DIN (0.04 ± 0.01 vs. 0.11 ± 0.01 ng/mL). Among heifers in mid diestrus on the day of PGF2α treatment, CLO reduced the interval to estrus compared with DIN (72.0 ± 2.2 vs. 82.4 ± 2.4 h). Prostaglandin F2α formulation and estrous detection treatment did not affect pregnancy to the first service. The interval between the first and second services tended to be reduced for the AED treatment compared with the MD treatment (24.4 ± 0.5 vs. 25.7 ± 0.6 d). Prostaglandin F2α formulation and estrous detection treatment did not affect the hazard of pregnancy. Although CLO treatment may shorten the interval to estrus in heifers at mid diestrus compared with DIN, PGF2α formulation did not affect reproductive performance. In the current experiment, no advantages in reproductive performance were observed when estrous detection was based on an AED compared with a MD.

Genomic merit for reproductive traits. II: Physiological responses of Holstein heifers.

Fertility traits were recently added to the evaluation of genetic merit, allowing for the selection of Holstein cattle with improved reproductive performance. In the current study, we investigated the associations among genomic merit for daughter pregnancy rate (GDPR) and heifer conception rate (GHCR) and physiological responses during proestrus and diestrus. Holstein heifers (n = 99) were classified based on GDPR [high = 3.26 ± 0.76 (1.6 to 5.3), n = 48; low = -0.17 ± 0.75 (-1.8 to 1.0), n = 51] and GHCR [high = 2.75 ± 0.77 (1.5 to 5.5), n = 49; low = 0.06 ± 0.67 (-2.1 to 1.2), n = 50]. Heifers were fitted with an automated estrous detection device, were treated with PGF2α for synchronization of estrus, and received either artificial insemination or embryo transfer at detected estrus. Blood was sampled at the time of PGF2α treatment, within 24 h of the onset of estrus (d 0), and on d 7, 14, 19 ± 2, 28, and 35. Blood samples from all heifers were analyzed for concentrations of estradiol (d 0) and progesterone (on the day of PGF2α treatment and d 0, 7, and 14). Blood samples from heifers pregnant on d 38 ± 3 were analyzed for concentrations of progesterone (d 0, 7, 14, 19 ± 2, 28, and 35), pregnancy-specific protein B (d 19 ± 2, 28, and 35), and insulin-like growth factor 1 (d 0, 7, 14, 19 ± 2, 28, and 35). Expression of mRNA for interferon-stimulated gene 15 in peripheral leukocytes isolated from blood collected on d 19 ± 2 was determined. Ovaries were scanned by ultrasound daily from d 0 to 4 or until ovulation was detected. Heifers with low GHCR tended to be less likely to be detected in estrus (78.0 vs. 91.8%). Estradiol concentration on d 0 was greater for heifers with high GDPR (4.53 ± 0.23 vs. 3.79 ± 0.23 pg/mL). The ovulatory follicle was larger for heifers with high GDPR (16.28 ± 0.33 vs. 14.55 ± 0.35 mm), whereas heifers with high GHCR tended to have smaller ovulatory follicles (15.00 ± 0.31 vs. 15.83 ± 0.37 mm). Heifers with high GDPR tended to be more likely to ovulate within 96 h of the onset of estrus (90.7 vs. 75.0%). Among heifers pregnant on d 38 ± 3, GDPR and GHCR were not associated with mRNA expression for interferon-stimulated gene 15. Heifers with high GDPR had greater concentration of pregnancy-specific protein B from d 28 to 35 (3.03 ± 0.15 vs. 2.48 ± 0.1 ng/mL). Heifers with high GHCR tended to have greater insulin-like growth factor 1 concentration from d 7 to 35 (108.0 ± 3.2 vs. 97.7 ± 4.2 ng/mL). Our results suggest that selection for Holstein cattle for GDPR may have positive effects on reproductive performance through changes in ovarian follicle development and steroidogenesis. Although selection of Holstein cattle for GHCR may negatively affect estrous expression by affecting ovarian follicle growth, selection for GHCR may improve reproductive performance by altering the somatotropic axis.

Low doses of bovine somatotropin enhance conceptus development and fertility in lactating dairy cows.

Objectives were to evaluate the effects of administering either one or two low doses of slow-release recombinant bovine somatotropin (bST) on hormone concentrations, conceptus development, and fertility in dairy cows. Cows from two farms were detected in estrus on or after 50 days postpartum (n = 1483), inseminated, and enrolled in the study (Day 0). Within farm, cows were blocked by parity and assigned randomly to receive a single placebo injection at insemination (control), a single injection with 325 mg of bST at insemination (S-bST), or two injections with 325 mg of bST administered on Days 0 and 14 (T-bST). From a subset of cows, blood was collected twice weekly from Day 0 to 42 for determination of hormone concentrations and on Day 19 for isolation of leucocytes and analysis of transcript abundance of selected interferon-stimulated genes. Pregnancy was diagnosed on Days 31 and 66, and ultrasonographic morphometry of the conceptus was performed on Days 34 and 48 in a subset of cows. Cows that received T-bST had increased plasma concentrations of GH and IGF1 for 4 wk, increased mRNA expression of ISG15 and RTP4 in leukocytes, earlier rise in the pregnancy-specific protein B in plasma of pregnant cows, increased conceptus size, and enhanced fertility. Cows that received S-bST had increased concentrations of GH and IGF1 for only 2 wk and it was insufficient to alter conceptus development and fertility. In conclusion, supplementation with low doses of bST during the pre- and peri-implantation periods enhanced conceptus development, reduced embryonic losses, and improved fertility in dairy cows.

Effects of fertility on gene expression and function of the bovine endometrium.

Infertility and subfertility are important and pervasive reproductive problems in both domestic animals and humans. The majority of embryonic loss occurs during the first three weeks of pregnancy in cattle and women due, in part, to inadequate endometrial receptivity for support of embryo implantation. To identify heifers of contrasting fertility, serial rounds of artificial insemination (AI) were conducted in 201 synchronized crossbred beef heifers. The heifers were then fertility classified based on number of pregnancies detected on day 35 in four AI opportunities. Heifers, classified as having high fertility, subfertility or infertility, were selected for further study. The fertility-classified heifers were superovulated and flushed, and the recovered embryos were graded and then transferred to synchronized recipients. Quantity of embryos recovered per flush, embryo quality, and subsequent recipient pregnancy rates did not differ by fertility classification. Two in vivo-produced bovine embryos (stage 4 or 5, grade 1 or 2) were then transferred into each heifer on day 7 post-estrus. Pregnancy rates were greater in high fertility than lower fertility heifers when heifers were used as embryo recipients. The reproductive tracts of the classified heifers were obtained on day 14 of the estrous cycle. No obvious morphological differences in reproductive tract structures and histology of the uterus were observed in the heifers. Microarray analysis revealed differences in the endometrial transcriptome based on fertility classification. A genome-wide association study, based on SNP genotyping, detected 7 moderate associations with fertility across 6 different chromosomes. Collectively, these studies support the idea that innate differences in uterine function underlie fertility and early pregnancy loss in ruminants. Cattle with defined early pregnancy success or loss is useful to elucidate the complex biological and genetic mechanisms governing endometrial receptivity and uterine competency for pregnancy.

Effects of resynchronization strategies for lactating Holstein cows on pattern of reinsemination, fertility, and economic outcome.

The objectives were to evaluate the pattern of re-insemination, pregnancy outcomes to re-insemination in estrus and at fixed time, and economic outcomes of lactating Holstein cows submitted to three resynchronization protocols. Cows were enrolled in the Experiment at 32 ± 3 d after pre-enrollment Artificial Insemination (AI), 7 d before pregnancy diagnosis, and randomly assigned to three resynchronization protocols. All cows diagnosed not pregnant at 39 ± 3 d after pre-enrollment AI were submitted to the Cosynch72 (Day 0 GnRH, Day 7 prostaglandin F(2α), and Day 10 GnRH and fixed time AI). Cows assigned to the control treatment received no further treatment, cows assigned to the GGPG treatment received a GnRH injection on Day -7, and cows assigned to the CIDR treatment received a controlled internal drug release (CIDR) insert containing 1.38 g of progesterone from Days 0-7. Cows observed in estrus were re-inseminated on the same day. Pregnancy was diagnosed at 39 ± 3 and 67 ± 3 d after re-insemination. Costs of the resynchronization protocols were calculated for individual cows enrolled in the study and pregnancies generated were given a value of $275. The GGPG treatment resulted in the slowest (P ≤ 0.06) rate of re-insemination. Overall pregnancy per AI (P/AI) at 39 ± 3 (P = 0.50) and 67 ± 3 (P = 0.49) d after re-insemination were not affected by treatment. Although cost of the control protocol was (P < 0.01) the smallest, return per cow resynchronized was (P < 0.01) greater for GGPG and CIDR protocols. We concluded that presynchronizing the estrous cycle of cows with GnRH or treating cows with a CIDR insert during resynchronization altered the pattern of re-insemination and improved the economic return of resynchronized cows.