Location
The study was carried out in a sheep farm – with the coordinates of 38.842174779662926, 37.28949932275392 – in Kalederesi Village of Gürün District of Sivas Province at an altitude of 1640.
Treatment Schedule
The material of this study has consisted of 140 Kangal sheep that became pregnant in autumn, gave birth in spring, and lost their lambs due to pneumonia and enteritis in about the first month (neonatal stage). At the beginning of the application, the average body weight was 44±5 kg, and the BCS (Body Condition Score) was between 2.5-3.25. The animals were from 55 to 70 days postpartum. The body condition scoring method explained by Ferguson et al. (1994) was used, and the BCS values of these ewes were scored from 1 to 5.
14 Kangal rams were also used to perform mating by sexual stimulation since the study was conducted in anestrus. When the farm records were examined, it was found that no pregnancy was achieved, although the sheep were with rams during anoestrous period in previous years. Progesterone (P4) measurements were made in the blood samples taken before the applications. The P4 value of animals in both groups was < 1 ng/ml. 140 sheep experiencing neonatal lamb losses were divided into two equal groups within each group. By starting synchronization 1 month before (-30th day), the animals were grazed only at nights for 45 days, and by keeping the animals in their barns during daylight hours, exposure to daylight was reduced. Thus, we waited for the postpartum physiological process during this period, including uterine involution, endometrial regeneration, and bacterial elimination of the uterus.
In the study, the animals in Group 1 (n:70) were administered an intravaginal sponge (20 mg fluorogestone acetate, Chronogest®, MSD) containing progesterone on day 0. 7 days after the application, the vaginal sponges were removed, and 263 µg PGF2α (d-cloprostenol, Estrumate®, MSD) and 600 IU equine chorionic gonadotropin (eCG) (Chorono-Gest/PMSG, Intervet International B.V., Boxmeer, Netherlands) were administered. The rams were introduced 1 day after the intravaginal sponge had been removed. On the 8th day of the study, the rams were separated from the herd. The animals in Group 2 were not manipulated, but they were just simultaneously introduced to the rams with the ones in Group 1 to benefit from the effect of ram.
Pregnancy examination
Following the rams were introduced, two pregnancy examinations were performed via trans-rectal on the 38th day and the transabdominal ultra-sonographic method on the 68th day. The trans-rectal and transabdominal examinations were achieved by using B-mode, linear-array ultrasonography device containing a 5.0-7.5 MHz rectal probe (Mindray DP50/Vet/US). The trans-rectal examinations were performed in the supine position. For the transabdominal examinations, the hairless area of the ventral abdominal wall just in front of the udder preferred for the placement of the probes. In the dorso-caudal direction of the udder, this area was thoroughly examined depending on whether pregnancy-related findings could be obtained or not. According to the gestation period in the ultrasonographic examination, after the detection of the gestational sac, the embryo/fetus, fluids, heartbeat, and placentomas, it was observed that the animal was pregnant. Lambing were followed and recorded.
Blood Analysis/assay
10 ml blood samples were taken from the vena jugularis to determine the progesterone level on the specified days of administration. After waiting for half an hour at room temperature, the blood samples were centrifuged at 3,000 revolutions/5 minutes in a cooled centrifuge, from each sample, blood was taken to Eppendorf tubes of 2 pieces of 1 ml and was stored at –80 °C until the measurement time. Progesterone measurement with analytical sensitivity ≤ 0.1 ng/ml and measurement range: 0.1 ng/ml-36.0 ng/ml progesterone was made by using the ARCHITECT Progesterone Chemiluminescence (7K77) Abbott assay and the fully automated ARCHITECT -i2000SR (Abbott) instrument via the Chemiluminescence Microparticle Immunoassay (CMIA) ) method. The intra-assay coefficient of variation (CV) ranged from 3.4-5.5% to 1.6-2.2% for low and high progesterone concentrations respectively. Analyses were validated for Serum (in serum and blood collected in serum separator tubes) and Plasma (with Na heparin, Li heparin, and K EDTA anticoagulants) samples. Validation was not performed with other anticoagulants other than the specified anticoagulants.
Method to be Used for Economic Analysis
Findings from the animals included in the study will be analyzed by using the formula below. As a result of the formula, it will be checked whether the method applied is "more economical" according to the findings obtained from the animals in the control group.
Economic Difference; EF = ∑ M1 - ∑ M2
Total Income of the Experimental Group; ∑ M1 = ( ∑ Kg1 - ∑P1 )
Total Income of the Control Group; ∑ M2 = ( ∑ Kg2 - ∑P2 )
EF: income differences
M1: The total economic income obtained as a result of the method applied to the animals in the experimental group
M2: Total income from animals in the control group
∑Kg: (Total lamb obtained x Market value*) / Current Dollar Rate**
∑P: (Applied Treatment Cost*** + Travel Fee + Veterinarian Practice Fee + Labour Cost****) / Current Dollar Rate
* The current market price for lambs is included in the calculation separately for males and females.
**(TCMB, 2021).
*** The costs such as Progesterone sponge + 500 IU (international unit) eCG + 263 µg PGF2α + the consumable costs (injectors, cotton, alcohol, etc.) per animal were included in the account.
.**** Calculation was made based on the total working hours over the minimum wage for 2020 (Aile Calisma, 2020).
Apart from this Calculation, that the economic value of the new lambs included may add to the farm will approximately be calculated using the formula below.
∑ MA* = Ad + Aey + Ds + Es + Ed
Ad: It was predicted that 80% of the female lambs could reach the ewe quality. The economic value of ewes was obtained by multiplying their number and market fair value.
Aey: It is predicted that the females who reach the brood-level can give birth to one lamb within the first year and two lambs in the other year, and half of those born will be male and the other half will be female. It is also predicted that all of the lambs obtained will be sold at the age of 6 months and they will provide an income for the farm. The income value is obtained by multiplying the market sales value of the female and male animals in this category with the number of the lambs sold. The income values of the currently born ones were ignored while calculating Aey.
Ds: It is predicted that 20% of the female lambs will be sold at the age of 6 months. Its economic value is obtained by multiplying the number of the sold ones and the market fair value.
Es: It is predicted that 80% of the males will be sold to the market. Its economic value is obtained by multiplying the males sold and the market fair value.
Ed: It is predicted that 20% of the males can reach breeding qualification. The economic value of breeding males was obtained by multiplying them and market fair value.
* While calculating the value of A, the costs of all animals included in the formula (such as feeding, maintenance, electricity, water, veterinary/health) are calculated and subtracted from the economic value.
Statistical analysis
Statistical analysis of all the data obtained in the study was evaluated using SPPS 25 (IBM Corp. Released 2017, IBM SPSS Statistics for Windows, Version 25.0. Armonk, NY: IBM Corp.) statistical package program. First, it was checked whether the variables met the assumptions of normality and homogeneity with Shapiro Wilk and Levene tests, respectively. The single effect of the treatments on oestrous rate, pregnancy, fertility, and multiple pregnancy rates were analyzed using the chi-square test. Fisher's exact test was used when one of the expected cell values was less than 5. Litter size data were log-transformed to attain normal distribution and analyzed with the independent sample t-test. In all analyses, P<0.05 was considered statistically significant.