Duration: 10/01/2002 to 09/30/2007

Administrative Advisor(s):

NIFA Reps:

Non-Technical Summary

Statement of Issues and Justification

Reproductive efficiency of cattle operations is suboptimal primarily due to: 1) failure of estrous cycles to resume after parturition, 2) poor fertility, and 3) poor detection of estrus in operations that use artificial insemination (AI). In attempts to improve reproductive efficiency, systems have been developed for programmed breeding. The recent research by the NC-1006 project has been instrumental in designing, testing, and optimizing systems that can be used for programmed artificial insemination (AI) in dairy and beef cattle herds. Many aspects of these reproductive management systems need to be optimized with basic research addressing mechanisms that reduce fertility in cattle and applied research to maximize economic benefits to livestock producers using available resources (i.e., FDA-approved hormones, labor, and facilities). Two major issues remain a focus for NC-1006 investigators and these will be addressed in studies during the next 5 years of research. 1) Recent research has demonstrated that anovulation is a surprisingly large problem in lactating dairy cows. For example, in the last joint project on lactating dairy cows by the NC-1006 committee, we found that 26.7% of lactating dairy cows were anovulatory following the voluntary waiting period. Recent studies by NC-1006 researchers that were discussed at our most recent technical committee meeting have continued to confirm a similarly high percentage of anovulatory cows based on multiple blood samples and multiple ultrasound evaluations of the ovaries. Thus, it seems clear that anovulation is becoming a more significant problem in lactating dairy cows. Modifying the Ovsynch protocol to synchronize the time of ovulation in lactating cows may substantially further reduce labor inputs for reproductive management; however, it may be possible to improve fertility with this treatment (one of the purposes of the current proposal). 2) In many beef cattle operations as many as 40 to 60% of cattle are anestrus at the beginning of the breeding season. None of the conventional programs prior to 1992 were designed to induce normal estrous cycles in anestrous cows. Adding a progesterone insert at the time of the GnRH injection improves fertility, especially in cows not cycling at the onset of the breeding season (contribution of the current NC-1006 project). Further refinement and success of these treatments should increase the convenience and appeal of applying AI to suckled beef cows. The current proposal, will test new treatments that are designed to synchronize ovulation in cycling cows and induce a fertile ovulation in anestrous cows and to determine the effects of a progesterone insert on embryonic survival.

JUSTIFICATION:

The Food Animal Integrated Research Symposium (FAIR 95) identified the need to "increase efficiencies of producing food from animals as a primary objective for future animal research." One of the key research areas with this objective was to "improve scientific understanding of reproductive mechanisms." Since FAIR 95, FAIR 2002 was convened as the second national conclave to establish consensus on animal agriculture research and education priorities for the 21st century. Six goals were established for FAIR 2002, each with its own set of objectives. The first of those goals was to "strengthen global competitiveness" and develop systems to keep U.S. animal agriculture competitive for the 21st century. Objective 1 of that goal is to "enhance production efficiency and economic strategies at the farm and ranch level." To achieve partly this objective, the report indicates that "research on different farming systems is needed to achieve efficiencies in reproduction." The NC-1006 committee is a long-standing group that has contributed greatly to the increase in reproductive efficiency of cattle since its inception in the early 1970's. Its long-term goal has been and continues to be consistent with the recent consensus goals set forth by FAIR 2002.

Our project encompasses the breadbasket states of the north-central region of the U.S. Included from that region in the current NC-1006 project are representatives from the Agricultural Experiment Stations of Iowa, Indiana, Illinois, Kansas, Michigan, Missouri, Minnesota, Ohio, and Wisconsin. Table 1 illustrates statistical information regarding the significance of the dairy and beef cattle industries in those nine states. It is clear that the numbers of dairy and beef cattle and their products, milk and meat, produced in the north-central region contribute a significant proportion (26-48%) of total production of the U.S. dairy and beef industries. Commodity receipts from sales of cattle and calves plus dairy products in the north-central region are significant, amounting to nearly $16 billion.
Reproductive inefficiency is one of the most costly and production-limiting problems facing both the dairy and cow-calf industries. For example, in the six mid-western states that comprise the Heart of America Dairy Herd Improvement Association, 17.4% of the dairy cows in 1999 were culled because of reproductive failure. It was only exceeded by death (18.2%) as the leading cause of culling (1999 Heart of America Annual DHIA Summary). Similar statistics are reported by other DHIA organizations in the U.S.

Cull beef cow sales makeup 15-20% of cow-calf herd total income (NAHMS, 1997). Over 27% of beef cows are culled because of reproductive failure or reproductive problems. Based on numbers of operations, that percentage is 34% of all cull cows. Further, these percentages increase with herd size: <50 cows (16%), 50-99 cows (30%), 100-299 cows (51%), and ƒ.ƒr300 cows (69%). Only the western region (46%) has greater culling for reproductive failure than the north-central region (37%). The NAHMS survey concluded that "by optimizing reproductive performance, producers can decrease forced culls" (i.e., cows sold for involuntary reasons).

Losses that occur because of reproductive failure are partly due to mismanagement of resources and the lack of adoption of appropriate technologies to sustain greater reproductive efficiencies. In the 1997 NAHMS survey of beef cattle producers, questions were posed regarding use of various reproductive technologies on ranches. Of the 1,190 producers surveyed, AI was used by only 13.3% of the operations; pregnancy palpation by only 34.5%; and estrus synchronization by 11.9% of operations. Of interest to our NC-1006 committee!&s long-term objectives were the reasons why estrus synchronization was not employed on those cow-calf operations surveyed. Their reasons were: "it doesn't work"(2%), "time and labor issues"( (36%), "lack appropriate facilities" (8%), "cost"(13%), "too complicated" (20%), and "other" reasons (21%). It was likewise interesting that those similar reasons were given, almost to the percentage point, for why those operations had not adopted the use of AI.

During the last 10 years, the NC-1006 project has contributed greatly to the development of several breeding programs to maximize pregnancy rates (Objective 1 of the current 1997-2002 project). These successes directly address the objections or reasons given by cow-calf producers for not adopting reproductive technologies. Information accrued by our group led to the development of the Ovsynch protocol (Pursley et al., 1995; 1997a,b; 1998) and its variations (i.e., the Cosynch protocol, progestins + the Ovsynch protocol) used in beef cattle (Thompson et al., 1999; Stevenson et al., 2000). These protocols generally increased pregnancy rates beyond controls because in both dairy and beef cattle, they induce fertile estrus and ovulation in postpartum cows that have not resumed estrous cycles by the end of the volunteer waiting period (dairy cows) or at the onset of the breeding season (beef cows).

During the past 5 years, our group conducted two cooperative experiments. The first experiment involved testing the efficacy of adding progesterone (via a progesterone-releasing intravaginal insert; CIDR-B; InterAg, Hamilton, NZ) to the Ovsynch protocol in lactating dairy cows. This experiment was replicated at seven (IL, KS, IN, OH, MI, MO, and WI) of the nine experiment stations involved in NC-1006 (Pursley et al., 2001). Addition of the CIDR to the Ovsynch protocol increased pregnancy rates in cows that had not resumed estrous cycles before the onset of the protocol from 34.7% (n = 95) with the Ovsynch protocol alone to 55.2% (n = 87) with the protocol plus addition of the CIDR. Overall, pregnancy rates increased from 40.9% (n = 320) to 50.8% (n = 313), respectively. In addition, we have reported that pregnancy rates after the Ovsynch protocol are improved further when the protocol is applied to cows in their early luteal phase (Vasconcelos et al., 1999) or when estrous cycles of cows are presynchronized with one (Cartmill et al., 2001) or two (El-Zarkouny et al., 2001) injections of PGF2ƒp administered 14 days apart, with the second or only injection given 12 days before the onset of the Ovsynch protocol. In a second experiment a similar protocol in suckled beef cows was tested where the Cosynch protocol was compared to the Cosynch protocol + supplemental progesterone provided by the CIDR (Lamb et al., 2001). This experiment was replicated at four (IL, KS, MN, and MO) of the seven stations. We found that pregnancy rates were increased from 48% (n = 188) with Cosynch protocol alone to 58% (n = 177) with the same protocol plus the CIDR. Both of these protocols were carried out without any estrus detection and all cows were inseminated at one fixed time.

Related, Current and Previous Work

Optimal reproductive management programs should maximize pregnancy rate (percentage of animals pregnant during the program) while minimizing costs (labor, hormones, etc.). Pregnancy rates are determined by multiplying the service rate (estrus detection rate) by the pregnancy rate per AI (conception rate). These 2 rates are clearly suboptimal in many cattle operations (Britt, 1985; Sanderson and Gay, 1996).

Dairy cattle.

Pregnancy rate per AI in high producing lactating dairy cows are less than desired. Pregnancy rates per AI have decreased from 66% in 1951, to about 50% in 1975, to less than 40% today (Aschbacher et al., 1956; Spalding et al. 1975; Butler et al., 1989; Pursley et al., 1997a, 1997b). However, pregnancy per AI in heifers has remained about 70% (Spalding et al., 1975; Hafs and Manns, 1975; Foote et al., 1979; Lee et al., 1983; Pursley et al., 1997a). The decrease in conception rates in lactating dairy cows is most likely related to the dramatic increase in milk production per cow during the same period (Oltenacu et al., 1980; Nebel and McGilliard, 1993). The mechanism by which milk production might decrease fertility is unknown.

One possible mechanism for decreased fertility is changes in circulating reproductive hormone concentrations particularly progesterone. Two types of studies indicate that low progesterone concentrations prior to AI result in lower fertility. Characterization studies (Fonseca et al., 1983; Rosenberg et al., 1990; Folman et al., 1990) have indicated a high correlation between plasma concentrations of progesterone in the luteal phase prior to AI and subsequent conception rates. For example Fonseca et al. (1983) showed an average of a 12.4% decrease in pregnancy rate per AI related to every 1 ng/ml decrease in progesterone concentration during the last half of the estrous cycle preceding first service. These studies clearly identified a physiologic phenomena, but did not define the mechanism causing this change in fertility.

A number of recent manipulative studies (Sirois and Fortune, 1990; Kinder et al., 1996; Savio et al., 1993; NE-161, 1996) used various methods to produce low plasma progestin concentrations and found aberrant growth of the dominant follicle. Low progesterone concentrations allowed extended growth of a dominant follicle causing what has been termed a persistent follicle. It is clear that conditions that produce a persistent follicle result in low fertility (Sanchez et al., 1993; Savio et al., 1993; Wehrman et al., 1993; Kinder et al., 1996; NE-161, 1996). For example, in a recent publication by the NE161 project the production of a persistent follicle decreased pregnancy rate per AI from 54% to 15% in lactating dairy cows (NE-161, 1996). It is likely that the decrease in fertility is due to a direct effect of increased LH pulsatility causing premature activation of the oocyte (Revah and Butler, 1996). However, a uterine effect cannot be discounted because Shaham-Albalancy et al. (1996) recently provided evidence that elevated progesterone during the estrous cycle may depress uterine secretion of PGFM and alter uterine endometrial morphology during the subsequent estrous cycle.

Progesterone supplementation of dairy cows for 7 days prior to a second injection of PFG2a increased pregnancy rates per AI compared to non-supplemented controls (Folman et al., 1990; Wehrman et al., 1993). In addition, high endogenous progesterone concentrations provided by a corpus luteum increased pregnancy rates during synchronization with norgestomet (Sanchez et al, 1993). A recent well-controlled experiment used a large number of dairy cows in New Zealand and found a slightly reduced pregnancy rate per AI after PGF2a treatment (Xu et al., 1996). The reduction was due to lower pregnancy rates per AI in cows bred after regressing a younger corpus luteum (d 5 to 8 of the estrous cycle) such that the ovulatory follicle would have developed in a lower progesterone environment (Xu et al., 1996). Supplementation with progesterone for 5 days prior to the PGF2a treatment using a controlled internal drug releasing device (CIDR) increased pregnancy rates per AI back to normal in cows in this early luteal phase (Xu et al., 1996). In contrast, Smith and Stevenson (1995, KS) observed similar conception rates in lactating cows and virgin heifers when either the PRID or a progestin (norgestomet) implant was used in a similar design, but only in cows that had a functional corpus luteum during progestin treatment. In cows without a corpus luteum, conception rates were less than controls; and particularly less after norgestomet than after PRID treatment. Norgestomet implants produce only a low progestin level and at least 2 implants are required to reduce LH pulsatility to levels observed during the mid-luteal phase (Sanchez et al., 1995). Our first proposed project will use a CIDR containing progesterone to increase serum progesterone concentrations and evaluate the effect on pregnancy rate per AI during a novel synchronization protocol, Ovsynch, that was developed as part of our previous NC-1006 project.

When GnRH was administered at random stages of the estrous cycle, the dominate follicle in 83% of lactating dairy cows and 45% of heifers would ovulate with subsequent initiation of a new follicular wave (Pursley et al., 1995, WI). When PGF2a was given 7 d after GnRH, luteolysis occurred and the dominate follicle arising from a new wave of follicles that emerged after GnRH was capable of ovulation in response to a second GnRH injection given 48 h after PGF2a (Pursley et al., 1995). This treatment (GnRH + PFG2a + GnRH + timed AI), known as Ovsynch, has been adopted by many dairy producers as a method for programmed AI-breeding. Pregnancy rates per AI resulting from such a treatment were similar to those achieved in lactating dairy cows bred to a detected estrus (Pursley et al, 1994; Pursley et al., 1997a, NC-1006 joint project; Pursley et al., 1997b). In contrast, pregnancy rates per AI were lower in heifers after Ovsynch and timed AI compared to AI after detected estrus, as would be expected from the low synchronization rates achieved with this protocol in heifers (Pursley et al., 1995; Pursley et al., 1997a, NC-1006 joint project). Using this treatment, pregnancy rates per AI in two other studies (Stevenson et al., 1996; Burke et al., 1996) were less compared to cows inseminated at estrus, but overall pregnancy rates (proportion of cows assigned to treatment that conceived) were similar. The improvement in service rate and the elimination of dependence on detection of estrus (reduced labor and errors) have made this protocol attractive to dairy producers in spite of the lack of improvement in pregnancy rate per AI.

In out more recent NC-1006 joint project, there was variation in pregnancy rates per TAI (%), percentage cycling, and average days in milk (DIM) among stations. However, in noncycling cows receiving Ovsynch + CIDR had greater (P<0.05) overall pregnancy rates (55.2%) than noncycling cows that received Ovsynch (34.7%) on day 28 after TAI. In addition, synchronization rate was similar after the Ovsynch (83.1%) and Ovsynch + CIDR (82.3%) protocols. We concluded that a CIDR inserted during the Ovsynch protocol enhanced fertility in non-cycling cows, but appeared to be ineffective in enhancing fertility in cycling cows; therefore, we intend to further investigate the possibility of increasing fertility in anovulatory cows.

Beef Cattle.

Estrus-synchronization programs in beef cattle are designed to reduce the breeding season, increase weaning weights, and group cows and heifers so AI can be used more efficiently (Odde, 1990). Treatments involving single (Fonseca et al., 1980; Kesler et all., 1980, Illinois) or multiple injections of GnRH (Edwards et al., 1983; Short et al, 1990), norgestomet implants (Ramirez-Godinez et al, 1981, KS; Smith et al, 1987; Troxel et al. 1993, IL), and(or) feeding melengestrol acetate (Patterson et al, 1989), with or without temporary calf removal, were used to induce ovulation and(or) estrus in cycling or anestrous suckled cows. Norgestomet prevented the short estrous cycle that normally follows first pubertal ovulations in heifers (Gonzalez-Padilla et al, 1975). Short estrous cycles in suckled cows that follow early weaning (Ramirez-Godinez et al, 1981) or ovulation induced by GnRH (Troxel and Kesler, 1984) or hCG (Garcia-Winder et al, 1986) are also prevented by pretreatment with norgestomet. The first short estrous cycle in suckled cows prevented the continuation of pregnancy when fertilization occurred (Ramirez-Godinez et al., 1982, KS).

Various treatment procedures developed to either induce or synchronize ovulation in anestrous and estrus-cycling suckled cows by administering GnRH and PGF2a, with (Troxel et al., 1993, IL; Hoffman et al., 1996, KS) and without a progestin source (Twagiramungu et al., 1995), have resulted in pregnancy rates and sometime pregnancy rates per AI that exceeded controls. Use of Ovsynch in suckled beef cows with fixed-time AI at the time of the second GnRH injection (48 h after PGF2a) or 24 h after the second GnRH injection has produced promising results (Geary and Whittier [1996], unpublished results; Colorado State University). Similar treatments (GnRH + norgestomet + PGF2a or GnRH + PGF2a) in suckled beef cows results in rates of estrus detection and pregnancy that exceeded controls given two injections of PGF2a (Forbes et al., unpublished results, KS). These Ovsynch or Ovsynch-like treatments induce estrus and subsequent pregnancy rates per AI equal those of estrus-cycling cows bred after a detected estrus regardless of whether AI was after detected estrus or a one fixed time. The use of higher progesterone supplementation that is possible with the CIDR in place of the norgestomet implant has not been tested in this model. The second experiment of Objective 1 will evaluate the effect of Ovsynch with or without progesterone supplementation on pregnancy rate per AI in cycling and anestrous beef cattle.

Previous data demonstrated that the administration of PGF2a 5d before synchronization of estrus with Syncro-Mate B decreased calving rates (Kesler et al., 1996, IL). It was further demonstrated that this decrease in calving rate occurred only in cows treated with Syncro-Mate B during the first half of the estrous cycle. Although Syncro-Mate B has been effectively used to synchronize estrus in beef cattle, the mechanism by which corpora lutea regress to the estradiol valerate and norgestomet injection has not been elucidated (Kesler and Favero, 1996, IL). In addition, norgestomet implantation during early embryo development enhanced the establishment of pregnancy in trials in which pregnancy rate in the control females was low (Favero et al, 1993; Favero et al, 1995; Domatob et al, 1996).

In our NC-1006 joint project (Lamb et al., 2001) we compared Cosynch to a treatment where a progesterone (CIDR) was added between the first injection of GnRH and the injection of PGF2a. Pregnancy rates were greater (P < 0.05) for Cosynch+Progesterone (58%) than for Cosynch-treated (48%) cows. Cows that had follicles >12 mm on day 2 had greater (P < 0.01) pregnancy rates than those with follicles less than or equal to 12 mm regardless of treatment. Pregnancy rates were similar between Cosynch and Cosynch+Progesterone treatments when cycling cows had elevated concentrations of progesterone at day 0, but pregnancy rates were greater (P < 0.05) in the Cosynch+P (79%) than in the Cosynch (43%) treatment when cycling cows had low concentrations of progesterone on day 0 (at PGF2a injection). Similarly, among noncycling cows, pregnancy rates were greater (P < 0.05) in the Cosynch+P (59%) treatment than in the Cosynch (39%) treatment. We concluded that treatment of suckled cows with Cosynch yielded acceptable pregnancy rates, but addition of a CIDR improved pregnancy rates in noncycling cows. Body condition and days postpartum at initiation of the breeding season affected overall efficacy of the Cosynch and Cosynch+P protocols.

Because the collection of categorical data requires large sample sizes for valid results, the NC-1006 group has collaborated well to provide results to be used in a typical production setting. With these thoughts in mind, the NC-1006 project will continue to test the effect of different progestin treatments on pregnancy rate and embryonic survival in beef cattle.

Objectives

1. Develop breeding protocols to maximize fertility in lactating cattle with special emphasis on anovulatory or anestrous cattle.
   
2. Determine the efficacy of using the CIDR to enhance pregnancy rates at a synchronized breeding in cattle and to resynchronize the first eligible estrus in non-pregnant cows after a previous insemination.
   

Methods

OBJECTIVES: Objective 1: Develop breeding protocols to maximize fertility in lactating cattle with special emphasis on anovulatory or anestrous cattle. Objective 1A. Identification and Treatment of Anovulatory Lactating Dairy Cows Rationale: Recent research has demonstrated that anovulation is a surprisingly large problem in lactating dairy cows. For example, in the last joint project on lactating dairy cows by the NC-1006 technical committee we found that 26.7% of lactating dairy cows were anovulatory. These data were based on collection of two blood samples 10 days apart. If both samples had low concentrations of progesterone, the cow was considered to be anovulatory. Recent studies by the NC-1006 researchers that were discussed at our most recent technical committee meeting have continued to find a similar high percentage of anovulatory cows based on multiple blood samples and multiple ultrasound evaluations of the ovaries. Thus, it seems clear that anovulation is becoming a more significant problem in lactating dairy cows. In order to effectively treat anovulatory dairy cows, methods to accurately identify these cows need to be developed and tested for practical use on commercial dairy operations. The standard identification method that has been utilized in research studies is to analyze multiple blood samples for circulating progesterone concentrations. Although progesterone analysis in milk can be practically performed, it has not become widely utilized. In addition, if a single time for progesterone evaluation could be identified this may make this method more practical for use in commercial operations. Studies have examined the efficacy of tail-head mounted heat detection devices, such as the Kamar, to identify the cows that are not showing estrus. These are placed on the tail head after calving and checked at the beginning of the breeding season to identify any non-cycling cows. The cows in these operations are on pasture most of the day and have much lower milk production than cattle in the U. S. It is unclear how accurate a similar strategy would be in identifying anovulatory cows in commercial dairy operations in the U.S. In addition, a single ultrasound evaluation at a critical time could be used to accurately distinguish anovulatory from ovulatory cows and this could be practically utilized in commercial dairy operations. The present study will utilize these three methodologies to determine the usefulness of each method in identifying non-cycling dairy cows in commercial operations. This study will be very large in order to provide potential differences among dairy operations in terms of incidence of anovulation as well as usefulness of each test in predicting anovulation. It is essential that many combine their expertise and data in order to provide a resource for commercial dairy producers on methods that could be practically utilized to identify anovulatory cows on their operations. A second rationale for evaluating a large number of cows with fairly intensive measurements is to provide more information on the types of anovulation that are occurring in dairy operations and correlate various physiological and management measures with anovulation. Recent data from one of our laboratories (Gumen and Wiltbank, unpublished) indicate that over 50% of anovulatory cows in dairy operations do not fit into any previously utilized definition for anovulation. Less than 20% of anovulatory cows were found to have follicular cysts (follicles >25 mm in diameter). Similarly, only about 20% of cows had follicles that grew to a maximal size that were less than that of a normal preovulatory follicle (>15 mm). Most of the anovulatory cows had follicles larger than ovulatory size but less than the classical size of follicular cysts. These cows would be extremely difficult to identify by the normal veterinary evaluations such as rectal palpation. Each experiment station contributing to the NC-1006 technical committee has personnel with expertise in performing ultrasound evaluations of the ovary. This expertise has been developed in previous joint projects that have been performed as part of this multi-state project. This expertise will now be focused on understanding anovulation in dairy cows from a perspective of incidence and causes. Normal epidemiology using only on-farm records cannot be utilized to accurately identify the type of anovulation that is occurring in individual cows (follicles that are less than or greater than ovulatory size or follicular cysts). Each of these anovulatory conditions may have different causes and incidences. In this study we will identify the type of anovulation using hormonal concentrations and ultrasound evaluation of follicular sizes. These data will be combined with a number of other measures such as body condition, disease incidence, and production parameters to provide correlative information on factors that may be associated with each type of anovulation using a large number of dairy cows and multiple commercial dairy operations. Obviously it is only useful to identify anovulatory cows on commercial operations if they can be effectively treated. In our previous joint project we found that treatment of cows with a CIDR (Controlled internal drug release; InterAg, Hamilton, NZ) insert in conjunction with the Ovsynch protocol increased pregnancy rates per AI (35% with Ovsynch vs. 55% with Ovsynch + CIDR; P < 0.01). In contrast, cycling cows did not benefit from combining a CIDR with the Ovsynch protocol (44% vs. 49% - P > 0.05). However, some differences among stations could not be easily explained. In this study we will analyze a much larger data set in which we have a great deal more information about the type of anovulatory condition in individual cows. This will allow us to evaluate whether addition of CIDR is beneficial for all certain types of anovulation. For example, the CIDR may be most effective in improving conception rates in cows with follicular cysts or cows with follicle growth to less than ovulatory size. It is critical to obtain large numbers of well-characterized anovulatory cows to understand the type of anovulatory condition(s) that are best treated with the CIDR. Thus, it is essential that multiple researchers combine data that is collected in an identical manner from multiple farms. This is clearly the advantage and mission of multi-state research projects and has been the strength of the NC-1006 project in the past. Hypothesis # 1: Anovulatory cows can be accurately identified by using a tail-head mounted heat detection device, ultrasound evaluation of ovaries, or milk progesterone analysis at the time of the first GnRH injection of the Ovsynch procedure. Hypothesis # 2: There will be significant associations between the different anovulatory conditions and the changes in body condition score, disease incidence, and milk production. Hypothesis # 3: Anovulatory cows with follicles greater than ovulatory size (>15 mm in diameter) will have an increase in conception rate after treatment with the CIDR. Experimental Protocol: There will be 7 experiment stations involved in this experiment and each station will work with one commercial dairy. Each station [IL, IN, KS, MI, MN, OH, WI] will evaluate 200 cows on the dairy for a total of 1400 cows. At calving, body condition score, calving difficulty, and any disease incidence will be recorded. At 12-19 days after calving a blood sample will be collected for analysis of concentrations of circulating non-esterified fatty acids (NEFAs) and glucose to help in determination of the metabolic state of the cow. All cows will be treated with the Presynch protocol starting at 40 days postpartum. Any cow at 40 or more days after calving (days 40-46 postpartum) (Lutalyse,awill receive 25 mg of prostaglandin F2 Pharmacia Animal Health). All cows also will receive a tail-mounted heat detector (either Kamar or Bovine Beacon). Fourteen days later treatment (25a(days 54-60 postpartum) cows will be given a second PGF2 mg). Fourteen days later the Ovsynch procedure will begin with GnRH treatment (25 mg),atreatment (100 5g), followed 7 days later by PG F2 followed 2 days later by GnRH treatment. (TAI) will be performed 10-18 hr after the second GnRH treatment. The inseminator will be blind to treatments of cows. A blood sample will be collected before each hormone treatment and at 5 and 12 days after the second GnRH treatment. The blood samples will be analyzed for circulating progesterone concentrations. Cows will have the ovaries evaluated by trans-rectal ultrasound at the time of the treatment and at the time of the first GnRH treatment. Inasecond PG F2 cows that have been found to be anovulatory, all samples will be analyzed for circulating estradiol concentrations to help with classification of anovulatory condition not only by maximal size of the follicle but also by high or low circulating estradiol concentrations. At 32 days after AI all cows will be evaluated for pregnancy using ultrasound. Fluid in the uterine horn will be evaluated and a heartbeat will be used as the definitive sign of a viable pregnancy. A second pregnancy evaluation will be done at 60 days to determine pregnancy loss. Calving data (gender of calf, twins, gestation length) will be recorded and analyzed. 1B. Efficacy of the CIDR in Enhancing Pregnancy Rates to a Synchronized Breeding and Synchronizing the Return Estrus for Non-pregnant Cows Two studies will be conducted under this objective. The objectives of the studies are to determine if administration of the CIDR after insemination 1) enhances the establishment of pregnant and 2) synchronizes estrus in cows that dont conceive to the synchronized insemination without compromising fertility. The first study will include 1,400 dairy cows from seven stations [IL, WI, OH, KS, MI, MN, and IN]. Each station will include 200 lactating dairy cows treated during the months of September to May to avoid summer heat. Cows will be assigned randomly to four groups: 1) untreated; 2) CIDR administration on days 5 to 14; 3) CIDR administration on days 14 to 21; and 4) CIDR administration on days 5 to 21. Before assignment to treatments, all cows will be synchronized with the Ovsynch protocol (GnRH followed 7 days later with an injection and then a second injection of GnRH 2 days after the injection"of PGF2 ). Each cow in treatments 2 and 4 will be administered a CIDR 5"of PGF2 days after the timed AI. The CIDRs administered to cows in treatment 2 will be removed on day 14. Each cow in treatment 3 will be administered a CIDR 14 days after the timed AI. For cows in treatment 4, the CIDRs administered on day 5 will be removed on day 14 and a new CIDR will then be administered. CIDRs administered on day 14 to cows in treatments 3 and 4 will be removed on day 21. Blood samples will be collected 10 days before and immediately before administering the first injection of GnRH for Ovsynch synchronization. Additional blood samples will be collected on days 5, 14, and 21 before administration and/or removal of the CIDR. All blood samples will be assayed for progesterone. The two blood samples collected before Ovsynch will be used to classify the cows as anestrus or cycling. The subsequent three blood samples will be used to assess the effect of the CIDR on altering synthesis of progesterone by the corpus luteum. All cows will be observed for estrus on days 5 to 25 to assess the synchrony of the return to estrus. All cows will be examined for pregnancy via transrectal ultrasound examination of the reproductive tract on days 28,42, and 56 after the TAI of the Ovsynch protocol. These examinations for pregnancy will permit not only the determination of pregnancy but embryonic loss up to 56 days post-insemination. Therefore, data (end points) collected from this study will include 1) pregnancy rate to the Ovsynch protocol; 2) synchrony and fertility at the return estrus; and 3) embryonic loss after the Ovsynch protocol. Other variables included in the analysis include milk production and days in milk.Only cows 50 to 250 days in milk with three or fewer insemination will be included in the study. The study design will permit evaluation of the effect of CIDR administration from 1) days 5 to 14; 2) days 14 to 21; and 3) days 5 to 21on efficacy of 1) enhancing the pregnancy rate to the Ovsynch protocol; 2) synchronizing the return estrus with normal fertility; and 3) reducing embryonic loss to the Ovsynch protocol. Preliminary data suggest that post-insemination progesterone supplementation via the CIDR may enhance the establishment of pregnancy. However, multi-station substantiation is necessary before recommendations can be made and it is unclear how supplemental progesterone affects pregnancy rate. Progesterone may improve pregnancy rates via greater early embryo development or reduced embryo loss. Resynchronization is the ability to synchronize the return estrus in cows that dont become pregnant to the first insemination with normal fertility. Again, preliminary data and data using the norgestomet implant, which is no longer available, demonstrate that supplemental progestins may effectively synchronize the return estrus with normal or less than normal fertility. However, again multi-station substantiation is necessary before recommendations can be made The second experiment will be conducted with the same objectives but with beef cows. This experiment will be designed as the previous experiment; however, only two transrectal ultrasound examination of the reproductive tract will be conducted (days 28 and 56 after the Ovsynch protocol). This study will include three stations [IL, KS, and MN] and 1,200 beef cows (400 cows per station). Objective 2: determine the efficacy of using the CIDR to enhance pregnancy rates at a synchronized breeding in cattle and to resynchronize the first eligible estrus in non-pregnant cows after a previous insemination. Objective 2A. The follicle from which it is derived influences developmental competence of the mammalian oocyte. We are just beginning to understand mechanisms occurring within the oocyte that impact development of the subsequent embryo. We hypothesize that in cattle, synchronization and superovulation regimens may affect not only ovarian dynamics but oocyte quality as well. In addition, implementation of these regimens must take oocyte viability into consideration as a major factor influencing subsequent embryo development, pregnancy rate and maintenance of pregnancy. We propose to examine the relationship of follicular development under the influence of the Ovsynch protocol on resultant oocyte and embryo quality [IN]. Objective 2B. Basic studies on pituitary cells will be performed using atomic force microscopy (AFM) to identify new cellular structures at the plasma membrane called 'pits' and 'depressions' where membrane-bound secretory vesicles dock and fuse to release vesicular contents. We will determine if such structures are present in neuroendocrine cells and whether these structures may be universal to secretory cells, where exocytosis occurs [IA]. Objective 2C. The relationship between reproduction and nutrition has been well documented. However, little is understood about the micro effects of nutrition on embryo quality and production or on follicle development in cattle during superovulation. There is dogma or speculation indicating that donor females consuming minerals from an organic source produces more embryos and those embryos have a higher quality than from donors consuming inorganic minerals. Therefore, we will determine effect of trace mineral nutrition on ovulation and embryo production parameters in superovulated females [MN]. Objective 2D. Reduced normal and abnormal follicular development are associated with the major objectives presented earlier in this proposal. In addition to development of new treatments to control follicular development, a better understanding of the basic mechanisms controlling follicular development will provide future insight to new improved treatment regimens. Studies will be conducted to examine the cellular and molecular mechanisms controlling ovarian follicular development [MO].

Measurement of Progress and Results

Outputs

• <b>Objective 1</b><ul> <li>At calving, body condition score, calving difficulty, and any disease incidence will be recorded. <li>At 12-19 days after calving a blood sample will be collected for analysis of concentrations of circulating non-esterified fatty acids (NEFAs) and glucose to help in determination of the metabolic state of the cow. <li>A blood sample will be collected before each hormone injection and at 5 and 12 days after the second GnRH treatment. The blood samples will be analyzed for circulating concentrations of progesterone. <li>Ovarian status of each cows will be evaluated by transrectal ultrasound at the time of the second PGF2&#61537; injection and at the time of the first GnRH injection. <li>In cows that have been found to be anovulatory, all serum samples will be analyzed for circulating estradiol concentrations to assist with classification of anovulatory condition not only by maximal size of the follicle but also by high or low circulating estradiol concentrations. <li>At 30-35 days after AI all cows will be evaluated for pregnancy using ultrasound. Detection of fluid in a uterine horn and(or) presence of a heartbeat will be used as the definitive sign of a viable pregnancy. <li>A second pregnancy evaluation will be carried out 4 weeks later to determine pregnancy loss. Calving results (gender of calf, twins, gestation length) will be recorded and analyzed.</ul> <li><b>Objective 2</b> <ul> <li>The two blood samples collected before Ovsynch will be used to classify the cows as anestrus or cycling. <li>The subsequent three blood samples will be used to assess the effect of the CIDR on altering synthesis of progesterone by the corpus luteum. <li>All cows will be observed for estrus on days 5 to 25 to assess the synchrony of the return to estrus. <li>All cows will be examined for pregnancy via transrectal ultrasound examination of the reproductive tract on days 28,42, and 56 after the TAI of the Ovsynch protocol. These examinations for pregnancy will permit not only the determination of pregnancy but embryonic loss up to 56 days post-insemination. <li>Therefore, data (end points) collected from this study will include 1) pregnancy rate after the Ovsynch protocol; 2) synchrony and fertility at the returned estrus; and 3) embryonic loss after the Ovsynch protocol. <li>Other variables included in the analysis include milk production and days in milk. Only cows 50 to 250 days in milk with three or fewer insemination will be included in the study. <li>The study design will permit evaluation of the effect of CIDR administration from 1) days 5 to 14; 2) days 14 to 21; and 3) days 5 to 21on efficacy of 1) enhancing the pregnancy rate to the Ovsynch protocol; 2) synchronizing the return estrus with normal fertility; and 3) reducing embryonic loss to the Ovsynch protocol.</ul>

Outcomes or Projected Impacts

• Our past two NC-113 regional research projects have focused upon timed breeding regimens in cattle. The Ovsynch protocol (1992-1997) project was adapted by many U.S. dairy producers and variations of Ovsynch (Select Synch and Cosynch) also have been adapted by both beef and dairy producers. In the last NC-113 renewal (1997-2002), multi-state studies using dairy and beef cows were conducted using the addition of CIDR inserts containing progesterone to the Ovsynch program. The rationale was that the CIDRs would supply additional progesterone and increase pregnancy rates in beef and dairy herds. Pregnancy rates were increased with the use of CIDRs in the Ovsynch protocol, with the biggest increase in pregnancy rates occurring in cows that were anestrus prior to treatment. Although the previous multi-state NC-113 projects have received considerable national attention and acceptance by producers, it is the aim of the current project to improve fertility and ease of adoption of the tested protocols. The multi-state studies planned in this project will provide new information on timing of treatments for breeding protocols, (e.g., the time from injection of PGF2&#61537; to injection of the second GnRH treatment) or the role of progesterone following breeding on conception rates and embryonic survival. One of the goals of the treatment regimens in beef herds is to make them more &#8220;user friendly&#8221; so that the AI will be more widely adopted. The task is to simplify protocols as much as possible and minimize handling and movement of cattle. It is also important that basic research of physiological mechanisms controlling the hypothalamic-hypophyseal-gonadal-uterine axis be obtained. A better understanding of the basic mechanism controlling reproductive function and fertility will provide the basis for development of new and better treatment regimens for controlling and programming reproductive function in cattle. Ultimately, fertility of cattle herds will be increased, and development of &#8220;user-friendly&#8221; breeding protocols will lead to increased adoption and use of AI by producers. As a result, U.S. cattle producers will be more competitive on the world market.

Milestones

(0):tion is a surprisingly growing problem in lactating dairy cows and suckled beef cows. For example, in the last joint project on lactating dairy cows by the NC-113 committee, we found that 26.7% of lactating dairy cows were anovulatory. In the last joint project on suckled beef cows by the NC-113 technical committee we found that 24% of beef cows were anestrus. In addition, reports have indicated that after fertilization up to 30% of pregnancies fail to be maintained to full term. A majority of these pregnancies are lost before or around the time of luteolysis, but recent evidence indicates that losses of 10 to 20% occurafter day 30 of pregnancy in suckled cows. Thus, it seems clear that anovulation is becoming a more significant problem in lactating dairy cows.

(0):ctively treat anovulatory dairy cows, methods to accurately identify anestrous cows need to be developed and tested for practical use on commercial dairy operations. This study will be very large to provide potential differences among dairy operations in terms of incidence of anovulation as well as usefulness of each test in predicting anovulation. We will also identify the type of anovulation using hormonal concentrations and ultrasound evaluation of follicular sizes. These data will be combined with a number of other measures such as body condition, disease incidence, and production traits to provide correlative information on factors that may be associated with each type of anovulation using a large number of dairy cows on multiple commercial dairy operations.

(0):ly, it is only useful to identify anovulatory cows on commercial operations if they can be effectively treated. In our previous joint project, we found that treatment of cows with a CIDR insert in conjunction with the Ovsynch protocol increased pregnancy rates per AI. In contrast, cycling cows did not benefit from combining a CIDR with the Ovsynch protocol. However, some differences among stations could not be explained. In this study, we will analyze a much larger data set in which we have a great deal more information about the type of anovulatory condition in individual cows. This will allow us to evaluate whether addition of CIDR is beneficial for all certain types of anovulation. It is critical to obtain large numbers of well-characterized anovulatory cows to understand the type of anovulatory condition(s) that are best treated with the CIDR. Thus, it is essential that multiple researchers combine data that is collected in an identical manner from multiple farms. This is clearly the advantage and mission of multi-state research projects and has been the strength of the NC-113 project in the past. The following hypotheses will be tested

(0): beef production systems are more extensive in nature than dairy systems, many beef producers are unable to monitor cattle daily or confine their cattle daily for artificial insemination. Therefore, most estrus-synchronization systems tend to focus on the first postpartum AI. By utilizing a used CIDR device two NC-1006 researchers (KS, MN) have recently synchronized the second or return estrous for potential AI. This system needs to be refined to allow producers only 2 or 3 days during which they need to monitor their cattle for repeat estrus. We will analyze a large data set in which we have sufficient information to understand whether supplementation with progesterone enhances pregnancy rates and whether resynchronization of estrus is a viable option for AI systems in beef cows. It is essential to combine data in an identical manner from multiple operations. This is clearly the advantage and mission of multistate research projects and has been the strength of the NC-1006 project in the past.

(0):h previous multistate NC-113 projects have received considerable national attention and acceptance by producers, it is the aim of the current project to improve fertility and ease of adoption of the tested protocols. O

Projected Participation

View Appendix E: Participation

Outreach Plan

In addition to multi-state refereed publications and presentations at scientific meetings the NC-1006 technical committee has made a commitment to solving practical reproductive management problems that can be applied by cattle producers immediately. In this manner through extension and experiment station reports along with producer workshops we intend to disseminate our results annually.

In addition, the newly formed North Central Bovine Reproductive Task Force is composed of mostly scientists in the north central region with major extension appointments. Their objectives are to provide leadership and consistency in programming to the dairy and beef industries in the area of reproductive physiology and management through Cooperative Extension efforts in the upper Midwest. Three specialists in this task force are principle investigators and station representatives of NC-1006. Their efforts have been coordinated and correlated with our project during the last 3-4 years. The task force has become a key disseminator of information to veterinarians and allied industry members through non-refereed peer reviewed publications and reproduction workshops. The natural overlap between the Task Force and NC-1006 will be a major focus for information dissemination.

The NE-161 committee is addressing parallel work but their focus is more general than ours. They are addressing reproductive insufficiency by examining multiple mechanisms and functions. We have met with NE-161 in the past and have made contact with them indicating a desire to combine meetings at one central site at least once in the next 5 years. Therefore, we can share data and create a new avenue for data dissemination among scientists.

Organization/Governance

The technical committee shall consist of one officially designated representative from each participating agricultural experiment station. The technical committee will meet annually. CSREES will designate one nonvoting representative. Officers will be elected for a period of one year, consist of Chairman and Secretary, and will progress from Secretary to Chairman. Elections will be held at the annual meeting. Officers will compose the executive committee. The executive committee, together with administrative advisors, is authorized to function on behalf of the technical committee in all matters pertaining to the regional project requiring interim action.

The Chairman, in consultation with the administrative adviser, shall arrange the time and place of the meeting, notify technical committee members of the meeting site, and prepare the agenda. The Chairman is responsible for preparation of the annual report of the regional project. The Secretary will record and distribute minutes of the annual meeting. Subcommittees may be appointed by the Chairman as needed for specific assignments. The executive committee will be in charge of coordinating cooperative research trials.

Literature Cited

Cartmill, J.A., S.Z. El-Zarkouny, B.A. Hensley, G.C. Lamb, and J.S. Stevenson. 2001. Stage of cycle, incidence and timing of ovulation, and pregnancy rates in dairy cattle after three timed breeding protocols. J. Dairy Sci. 84:84:1051-1059.

El-Zarkouny, S.Z., J.A. Cartmill, A.M. Richardson, M.A. Medina-Britos, B.A. Hensley, and J.S. Stevenson. 2001. Presynchronization of estrous cycles in lactating dairy cows with Ovsynch + CIDR and resynchronization of repeat estrus using the CIDR. J. Dairy Sci. 84:(Suppl. 1):249 (Abstr. #1028).

Favero, R.J., D.B. Faulkner and D.J. Kesler. 1993. Norgestomet implants synchronize estrus and enhance fertility in beef heifers subsequent to a timed artificial insemination. J. Anim. Sci. 71:2594-2600.

Favero, R.J., D.B. Faulkner, T.G. Nash, and D.J. Kesler. 1995.Effect of norgestomet treatment after insemination on the calving rate of postpartum suckled beef cows. J. Anim. Sci. 73:3230-3234.

Lamb, G.C., J.S. Stevenson, D.J. Kesler, H.A. Garverick, D.R. Brown, and B.E. Salfen. 2001. Inclusion of an intravaginal progesterone insert plus GnRH and prostaglandin PGF2ƒp for ovulation control in postpartum suckled beef cows. J. Anim. Sci. 79:2253-2259.

NAHMS, 1997. http://www.aphis.usda.gov:80/vs/ceah/cahm/Beef-Cow-Calf/beef.htm.

Pursley, J.P., P.M. Fricke, H.A. Garverick, D.J. Kesler, J.S. Ottobre, J.S. Stevenson, and M.C. Wiltbank. 2001. Improved fertility in noncycling lactating dairy cows treated with progesterone during Ovsynch. Midwest ADSA/ASAS Mtg., Des Moines, IA, March19-21, (Abstr. #251).

Pursley, J. R., M. R. Kosorok, and M. C. Wiltbank. 1997a. Reproductive management of lactating dairy cows using synchronization of ovulation. J. Dairy Sci. 80:301-306.

Pursley, J. R., M. O. Mee, and M. C. Wiltbank. 1995. Synchronization of ovulation in dairy cows using PGF2a , and GnRH. Theriogenology 44:915-923.

Pursley, J. R., R.W. Silcox, and M. C. Wiltbank. 1998. Effect of time of artificial insemination on pregnancy rates, calving rates, pregnancy loss, and gender ratio after synchronization of ovulation in lactating dairy cows. J. Dairy Sci. 81:2139-2144.

Pursley, J. R., M. C. Wiltbank, J. S. Stevenson, J. S. Ottobre, H. A. Garverick, and L. L. Anderson. 1997b. Pregnancy rates per artificial insemination for cows and heifers inseminated at a synchronized ovulation or synchronized estrus. J. Dairy Sci. 80:295-300.

Stevenson, J.S., K.E. Thompson, W.L. Forbes, G.C. Lamb, D.G. Grieger, and L.R. Corah. 2000. Synchronizing estrus and(or) ovulation in beef cows after combinations of GnRH, norgestomet, and prostaglandin F2ƒp with or without timed insemination. J. Anim. Sci. 78:1747-1758.

Thompson, K.E., J.S. Stevenson, G.C. Lamb, G.M. Grieger, and C.A. Loest. 1999. Follicular, hormonal, and pregnancy responses of early postpartum suckled beef cows to GnRH, norgestomet, and prostaglandin F2ƒp. J. Anim. Sci. 77:1823-1832.

VanCleeff, J.V., K.L. Macmillan, M. Drost, M.C. Lucy, and W.W. Thatcher. 1996. Effects of administering progesterone at selected intervals after insemination of synchronized heifers on pregnancy rates and resynchronization of return to service. Theriogenology 46:1117-1130.

Vasconcelos, J.L.M., R. W. Silcox, G. J. Rosa, J. R. Pursley, and M. C. Wiltbank. 1999. Synchronization rate, size of the ovulatory follicle, and pregnancy rate after synchronization of ovulation beginning on different days of the estrous cycle in lactating dairy cows. Theriogenology 52:1067-1078.

Attachments

Land Grant Participating States/Institutions

FL, IA, IL, KS, KY, MI, MO, NE, OH, SD, WI

Non Land Grant Participating States/Institutions

USDA ARS