# Presentation: 254914

## Reproductive Physiology and Management Research Program

- Robert Dailey, Jorge Flores, Keith Inskeep, Marlon Knights, Paul Lewis and Matthew Wilson

## Projects/Support

- Hatch 427/NE1007 - Ovarian and Environmental Influences on Embryonic/Fetal Mortality in Ruminants 10/1/02-9/30/07
- State 149 (Animal Health Funds) - Effect of Peptidoglycan-Polysaccharide (PG) Complex of Streptococcal Cell Wall on Reproductive Efficiency and Mastitis in Sheep, 10/1/00 – 9/30/02 - Dailey
- State 170 (Animal Health Funds) – Pattern of Cytokines in Uterine and Vaginal Tissue and Secretions during Discrete Stages of the Estrous Cycle of Sheep, 10/1/02 – 9/30/04 - Dailey

## Projects/Support con’t.

- State 140 (WV Legislature/WV Department of Education)- WV Sheep Improvement Project – Increasing Farm Income by a Strategic Shift in Breeding Time for Ewe Flocks, 7/1/98 – 6/31/07 Lewis and Inskeep
- NRSP-7 – Effectiveness of Intravaginal Progesterone Inserts... and FSH ... Ewes, 1998-2003 - Inskeep, Lewis and Knights
- USDA-NRI 2001 -35203-10982 Investigation of the Benefits of Early Gestational IGF-I in Sheep, 9/1/01-8/31/04 – Wilson

## Support from industry and other sources

- Pharmacia Animal Health		Nat. Inst. Occup. Safety & Hlth.
- InterAg Division of DEC International	The Ohio State University
- Vetrepharm				Dept. of Pathology, W V U
- Elysian Fields Farm (PA)		WV Sheep Federation
- University of Michigan			Colorado State University
- ARS Forage Unit, Beaver, W V 		Impres Medical Corp.
- Intervet					Dept. of Immunology, W V U
- Dept. of Biology, W V U			Jeff Lawson Farm (VA)
- USDA Sheep Exp. Station, DuBois ID	Dale Evans Farm (MD)
- Allen Pysell Farm (MD)			Richard Blamble Farm (MD)
- Michigan State University 		University of Wyoming
- 20 W Va Sheep Producers		Antonio Lopez S. - Spain

## Philosophical Approaches

- Conduct basic research into factors that limit reproductive efficiency and management of sheep in West Virginia and the Appalachian Region.
- Conduct applied research to test concepts generated by basic research, to enable wide application of new concepts, and to identify other problems deserving basic studies.
- Collaborate to maximize understanding and output.

## Research Areas

- Regulation of Luteal Function
- Induction & Synchronization of Estrus
- Early Embryonic Mortality
- Late Embryonic and Fetal Mortality
- 	- Progesterone Metabolism
- Mastitis and Reproduction
- Embryonic and Postnatal Growth
- Regulation of Follicular Dynamics

## Regulation of Luteal Function

- Current and Proposed Studies:
- Changes in gene expression of the prostaglandin synthesis and endothelin systems, and of prostaglandin dehydrogenase activity in corpora lutea.
- Signal transduction mechanisms mediating actions of endothelin and prostaglandin in luteal cells:
- 		- Changes in concentration of intracellular 		Ca++
- 		- Role of protein kinase C isozymes, 			especially .

## Induction & Synchronization of Estrus

- Anestrous Ewes (Knights)
- Three studies evaluated the CIDR-G (to deliver progesterone) and single injections of FSH for out-of-season breeding.  One goal was to maximize conception rates.  The second goal was to increase ovulation rate and litter size (to 2.0).	(1392 ewes)
- Conclusions: Progesterone increased pregnancy rates to two services from 45% with ram introduction alone to 62%. Treatment for 5 days was as effective as treatment for 12 days. FSH marginally increased ovulation rate, but did not consistently increase litter size.
- These data have been reported to USDA-NRSP7 for submission to FDA for approval of the CIDR-G for anestrous ewes.  Three papers published.

## Induction & Synchronization of Estrus

- Ewes during the breeding season (Knights and Dixon)
- Three studies were conducted with CIDR-G to synchronize estrus during the breeding season. Treatment with 2 CIDR-G for 12 days yielded pregnancy rate to first service and prolificacy of 90% and 1.9 vs 79% and 2.0, respectively, (ns) in control ewes.  401 ewes
- Treatment with CIDR-G for 5 days with PGF2α 24 h before insert removal increased pregnancy rate to first service (63%) compared to ewes that received PGF2α only (44%; P < .05).  164 ewes
- Treatment with CIDR-G for 5 days with PGF2α at insert removal increased pregnancy rate to first and second services (71 and 72% P < .05) compared to ewes that received PGF2α only (58 and 61%; P<.01).  1014 ewes

## Breeding Anestrous Ewes Using Ram Effect and GnRH

- Ram is a powerful tool to induce ovulation in anestrous ewes.
- 	- It can be enhanced by injection of progesterone at ram introduction.
- 	- Estrus can be synchronized with subsequent injection of PGF2α (day 12-16), but pregnancy rates to two services usually do not exceed 70%.
- Objectives:  	Will more ewes ovulate if ram introduction is _followed_ by injection of GnRH?
- 		Will GnRH enhance pregnancy rates in response to the ram effect and PGF2α?
- In two trials to date, GnRH has not increased estrous response or pregnancy rate.  			(Jordan, Wurst and Knights)

## Induction & Synchronization of Estrus

- Ewe lambs during anestrus 	(Knights and Baptiste)
- Fall born ewe lambs were treated in July.  Groups were:
- 	Control
- 	Used CIDR-G device (progesterone)
- 	Used CIDR-G device and ram introduction
- 	 Used CIDR-G device, ram introduction and 25 g EB 24 h after
- 		CIDR withdrawal
- Ram introduction alone increased LH pulse frequency, follicular development and ovulation.  Progesterone alone did not.
- Progesterone and EB combined with ram introduction increased estrous response.

## Late Embryonic & Fetal Mortality

- Studies in Ewes :

## Anestrous Ewes That Experienced Pregnancy Loss From Day 25 to Lambing 

**Anestrous Ewes That Experienced ****Pregnancy Loss From Day 25 to Lambing**

- 1998

- 1999

- _________________________________________________

- Synchronization Studies - Knights

## Determining Late Embryonic and Fetal Loss

**Summer 2000 and 2001 (N = 1400)****Breeding   -  May to September**
**1.  Embryos / Fetuses on each day of pregnancy      ****	****diagnosis **
**2.  Lambs at term **
_**d25**__**	**__**	**__**45**__**	**__**	**__**65**__**	**__**	**__**85**__**	**__**	**__** Term**_
**        ****Loss****	****       Loss****	****       Loss****	****	****Loss**

**Determining Late Embryonic and Fetal Loss**
**in Ewes**

## Effect of Face Color On Embryo or Fetal Loss During Several Stages of Gestation 

- a, b,c Columns with different letters are different (P < 0.05)

## Ewes Losing Embryos Present at Day 25 of Gestation By Type of Loss

**Multiple  Partial**

**Single Complete**

**Multiple Complete**

## Corpora Lutea (Ovulations) Not Represented By Lambs Born in Ewes Treated With Three Dosages of FSH

- Knights et al., 2002

## Effect of Mastitis on Reproductive Efficiency

**Mastitis**

**Bacteria**

**Cell Wall Components (peptidoglycan, lipopolysaccharide)**

**Inflammatory               Mediators**

**Conception/**
**Pregnancy **
**Failure**

## Mastitis and Reproduction in Dairy cowsAmanda Barker – Tennessee data

- 				Mastitis Caused:
- Increased Number of Services
- Increased Interbreeding Interval
- Increased Days Open
- Increased Calving Interval

**Cell Wall of Gram-Negative and Gram-Positive Bacteria**

**    ****Hurlbert, 1999**

## Sheep Model for Effects of Mastitis on Reproduction      (Amanda Stewart)

**Rationale: **
**Bacteria promote inflammation and infection after the host digests the bacterial cell wall**
**Major component of cell wall is peptidoglycan (PTG) **
**Majority of embryonic loss in cows is by days 6- 8**
**Method:  Inject PTG in ewes on day 5 post-breeding**
**Results:  - Induced transient fever****	**
**	****	****    ****- Reduce progesterone****	****	****	****	****	****	****    - Increased cortisol****	****	****	****	****	****	****    - Increased prostaglandin metabolite (PGFM)**
**	****	****    ****- Decreased pregnancy rate**

## Effect of Blocking Inflammatory Increase in PGF2a on Pregnancy Rate          (Amanda Stewart)

**Rationale: ****PTG increased PGFM.  Did increased PGF****2******** **** decrease pregnancy rate?****  **
**Objective:**
**Does blocking PGF****2******** ****following PTG challenge result in increased embryo survival?**
**Are other inflammatory mediators involved?**
**Treatments:**
**30 ********g PTG/kg  body weight on day 5** plus saline (n = 10)
- PTG plus flunixin meglumine (fm) **at 0, 6, and 12 h** (n = 10)
- Saline plus fm (n = 4)
- Saline alone (n = 5)
- Jugular blood:** **Progesterone, Inflammatory mediators:  TNF-, IFN-, TGF-, IL-1, IL-2

## Results - Pregnancy Rates

- Peptidoglycan plus saline    	60%
- Peptidoglycan plus FM		50%
- Saline plus FM				75%
- Saline					80%
**Conclusion:**
**	****Increased PGF2a not solely responsible for pregnancy losses.**

## 
Primary objective was to determine if immunization prevented:
	1. fever at bacterial challenge 
	2. early embryonic loss, and	3. mastitis in ewes.

Five days after breeding, sheep were injected s.c. 		with PTG.

**Primary objective was to determine if immunization prevented:**
**	****1. fever at bacterial challenge **
**	****2. early embryonic loss, and****	****3. mastitis in ewes.**
**Five days after breeding, sheep were injected s.c. ****	****	****with PTG.**

**Effect of Peptidoglycan on Reproductive Efficiency and Mastitis in Sheep****      (Holásková)**

**Immunization did not: **
**	****prevent fever on day 5,**
**	****enhance pregnancy maintenance, **
**	****prevent mastitis after weaning.**
**In fact, increased dosage of PG-PS lowered ****	****	****	****the probability of successful pregnancy. **
**	****This result fits earlier finding by Coleman and Dailey ****	****that reproductive disorders occurred more ****	****commonly in cows that had previously experienced ****	****the disorder.**
**Mastitis tended to occur in fewer immunized ewes.**

**Results**