Intrauterine proteolytic enzymes therapy hastens expulsion of fetal membranes in dystocia affected buffaloes.

Background: Difficult calving (dystocia) in buffalo cows is a major obstetrical problem which further leads to metritis complex, encompassing the retention of fetal membranes (RFM), puerperal metritis, endometritis and pyometra with impaired future fertility. Aims: The current study aimed to evaluate the effect of the administration of intrauterine proteolytic enzymes on the expulsion of fetal membranes and postpartum fertility in dystociac buffaloes. Methods: Proteolytic enzymes consisting of Trypsin (16 mg), Chymotrypsin (16 mg), and Papain (8 mg) were dissolved in 500 ml normal saline were administered after 1 h of assisted delivery in dystociac buffaloes along with the conventional therapy. Results: The treated animals (n=15) expelled fetal membranes within a shorter period of time (P=0.043) compared to the control group (n=15) with none in the treatment group retaining it for more than 24 hours. Fewer (26.67 vs 73.33%; P=0.027) postpartum uterine infections developed in the treated animals compared to the control group. The interval between first postpartum estrus (P=0.067), service period (P=0.554), and open days (P=0.557) was shorter in the treatment group compared to the control group where postpartum anestrus developed less frequently (26.67 vs 66.67%; P=0.066) in the animals treated with enzymatic therapy. Systemic illness (neutrophillia) was reduced in the treatment group compared to the control on day 20 (64.55 ± 1.14% vs 70.23 ± 0.99%; P=0.001) and 45 (55.05 ± 1.63% vs 64.92 ± 1.45%; P<0.001) postpartum. Conclusion: It is concluded that proteolytic enzymes therapy after assisted delivery in dystociac buffalo cows could help in the early expulsion of fetal membranes and reduce uterine infections with decreased neutrophils count.

In vitro evaluation of Labrador dog spermatozoa cryopreserved in Tris-citric acid-fructose buffer supplemented with different combinations of extracellular and intracellular cryoprotectants.

Aim of this study was to compare different combinations of penetrating intracellular CPAs, i.e., glycerol (G), ethylene glycol (EG), propylene glycol (PG), dimethyl formamide (DM), and methyl acetamide (MA) and extracellular [egg yolk (EY), egg yolk plasma (EYP), low-density lipoproteins (LDL), and coconut water (CW)] in Tris-citric acid-fructose buffer (T) for Labrador dog semen cryopreservation. The study was conducted in two parts, first trial was conducted to assess optimum glycerol concentration (5-7%) in TEY and equilibration time (ET, 2-4 hrs) for Labrador dog semen cryopreservation. Secondly, compatibility of 15% TEY, 15% TEYP, 13% TLDL, and 25% TCW with G, DMF, MA, D + M, EG, and PG was evaluated for in vitro sperm function tests. Decline in sperm attributes, i.e., motility, viability, plasma membrane integrity (PMI), and acrosome integrity (AI)) was significantly (p < 0.05) less in 7% TEY-G and 4 h compared to other concentrations and ET at post-thaw. There was significantly (p < 0.05) less decline in sperm attributes in TEY-G, TEYP-G, TLDL-G, TLDL-D, TLDL-EG, and TCW-D extenders compared to other combinations at post-thaw. However, these parameters were significantly (p < 0.05) high in TEY-G and TEYP-G compared to TEYP-D, TLDL-G, TLDL-D, TLDL-EG, and TCW-D extenders at post-thaw. However, decline in motility, viability, PMI, and AI was identical in these seven extenders. This study concluded that glycerol at a concentration of 7% in TEY and 4 h ET were optimum for successful cryopreservation and besides TEY-G, other combinations of protectants may be an alternative for canine semen cryopreservation.

Subclinical endometritis in estrual buffaloes: diagnosis, prevalence and impact on reproductive performance.

The study evaluated the reliability of polymorphonuclear (PMN) cell count in endometrial cytology by cytobrush technique as a diagnostic for subclinical endometritis (SCE) at the time of estrus and its relationship with bacterial presence and fertility in buffaloes. Healthy pluriparous buffaloes (n = 115) were subjected to endometrial sampling by cytobrush technique (for cytology and bacterial isolation) and ultrasonography of genitalia prior to insemination at the time of spontaneous estrus. Buffaloes were inseminated at the same estrus and were confirmed for pregnancy by ultrasonography at day 40 post-insemination. Growth of aerobic bacteria was recorded in 26.1% (30/115) of estrual buffaloes. Bacteria of eight distinct genera (Staphylococcus spp., Bacillus spp., Proteus spp., Escherichia coli, Klebsiella spp., Streptococcus spp., Pseudomonas spp. and T. pyogenes) were isolated. Presence of bacterial infection was considered as the standard, against which PMN threshold for SCE was calculated by receiver operator curve (ROC). The PMN count of ≥ 5% with good sensitivity (66.7, 95% CI 50.9-81.4) and specificity (87.0%, 95% CI 78.0-94.6) with an overall high diagnostic accuracy (81.7%) for predicting SCE was obtained. Based on calculated PMN threshold, buffaloes were divided into subclinical endometritic (SCE; ≥ 5% PMNs) and control (< 5% PMNs) groups. Thirty-one buffaloes (27.0%, 31/115) had ≥ 5% PMNs in the endometrial cytology at estrus. Ultrasonographic evaluation showed that size of largest follicle, endometrial thickness, and uterine horn diameter did not differ significantly (P > 0.05) between the two groups. Significantly, lower conception rate (22.58 vs 36.90%, P < 0.05) for first artificial inseminations (AIs) and significantly higher number of AIs per pregnancy (2.81 ± 0.27 vs 1.75 ± 0.15, P < 0.05) were recorded in SCE compared to control group. The results indicated that cytobrush based endometrial cytology with 5% PMN cut-off value has a good accuracy for predicting subclinical endometritis at estrus in buffaloes.

Comparison of in vitro and in vivo fertilizing potential of buffalo bull semen frozen in egg yolk-, soya bean lecithin- and liposome-based extenders.

The objective of this study was to compare different extenders for post-thaw in vitro sperm function and in vivo fertility of buffalo semen. Accordingly, sperm of 30 ejaculates extended in egg yolk (TRIS with 20% egg yolk; EY), two soya lecithin-based (SL-1; AndroMed® and SL-2; Bioxcell® ) and a liposome-based extender (LS; OptiXcell® ) were tested. The post-thaw semen was evaluated for computer-assisted sperm analysis (CASA), sperm viability, membrane and acrosome integrity, DNA integrity and acrosome reaction and first service pregnancy rate (FSPR) in a fixed-time artificial insemination programme. Total motility and VCL were the only CASA-based parameters that exhibited significantly higher (p < .05) percentage in LS among these extenders. Post-thaw percentage of acrosome integrity (55.9 ± 1.4, 58.1 ± 2.0, 55.8 ± 2.0, 56.6 ± 2.3) and DNA integrity (68.8 ± 2.0, 69.2 ± 2.3, 71.3 ± 2.1, 69.1 ± 2.1) did not differ (p > .05) in EY, SL-1, SL-2 and LS extender, respectively. However, a variable response in terms of efficacy of different extenders for sperm viability and plasma membrane integrity was observed. Assessment of inducibility of acrosome reaction showed significant differences between extenders (51.9 ± 2.1, 44.3 ± 2.4, 46.1 ± 2.3 and 58.1 ± 3.1%, respectively, for EY, SL-1, SL-2 and LS). Furthermore, field trials revealed significantly higher (p < .05) FSPR of LS-extended semen as compared to that for EY, SL-1 and SL-2 extender (46.3%, 41.2%, 31.2% and 29.7%, respectively). It is concluded that the liposome-based extender is more effective than egg yolk- and soya lecithin-based extenders and may be used for cryopreservation of buffalo semen in the future.

Effects of preovulatory follicle size on estradiol concentrations, corpus luteum diameter, progesterone concentrations and subsequent pregnancy rate in buffalo cows (Bubalus bubalis).

The present study was undertaken to investigate the effects of preovulatory follicle (POF) size on estradiol concentrations, luteal profile (CL diameter and progesterone concentration) and subsequent pregnancy rate in Murrah buffalo cows. The buffalo cows (n = 49) were synchronized for estrus by two doses of PGF2α given 11 days apart. The buffalo cows were inseminated during standing estrus and again after 24 h. Ovaries were scanned at estrus and 24 h intervals until ovulation, thereafter on days 5, 12 and 16 post-ovulation to examine the POF and CL diameter. Size of POF at estrus was divided into three categories; I: 10 to ≤12; II: >12.0 to ≤14.0; III: >14.0-16.0 mm. Blood samples were collected for estradiol (on day of estrus) and progesterone concentration (on days 5, 12 and 16). The estradiol concentrations were greater (P < 0.05) in category II than category I with the greatest (P < 0.05) concentrations estimated in category III. A positive correlation (P < 0.05) between POF and progesterone concentration, CL diameter and progesterone concentration was observed on all sampling day. Pregnant buffalo cows exhibited greater (P < 0.05) plasma progesterone as compared with their non-pregnant counterpart. Greater pregnancy rates were observed with an increased size of POF (χ2 = 2.9, P > 0.05). It was concluded that the POFs having diameters between 12 and 16 mm are mature enough to be transformed into CL of such optimum diameter and can secrete optimum progesterone concentrations that can sustain the pregnancy in Murrah buffalo cows.

Effect of superstimulation protocols on nuclear maturation and distribution of lipid droplets in bovine oocytes.

Our objective was to study the effect of superstimulation protocols on nuclear maturation of the oocyte and the distribution of lipid droplets in the ooplasm. Heifers (n=4 each group) during the luteal phase were either treated with FSH for 4 days (Short FSH), FSH for 4 days followed by 84h of gonadotropin free period (FSH Starvation) or for 7 days (Long FSH) starting from the day of wave emergence. In all groups, LH was given 24h after induced luteolysis (penultimate day of FSH) and cumulus-oocyte complexes were collected 24h later. Oocytes were stained for nuclear maturation (Lamin/chromatin) and lipid droplets (Nile red). The Long FSH group had a greater proportion of mature oocytes (metaphase II) compared with heifers in the Short FSH and FSH Starvation groups (59/100 vs 5/23 and 2/25, respectively; P<0.01). On average across all groups, oocytes contained 22pL of lipids (3.3% of ooplasm volume) distributed as 3000 droplets. Average volume of individual lipid droplets was higher in the FSH Starvation (11.5±1.5 10(-3) pL, P=0.03) compared with the Short and Long FSH groups (7.2±0.6 10(-3) and 8.0±0.8 10(-3) pL, respectively). In conclusion, both FSH Starvation and Short FSH treatments yielded a lower proportion of mature oocytes compared with the Long FSH treatment. Furthermore, FSH starvation led to an accumulation of larger lipid droplets in the ooplasm, indicating atresia. Our results indicate that a longer superstimulation period in beef cattle yields higher numbers and better-quality oocytes.

Comparison of follicular dynamics, superovulatory response, and embryo recovery between estradiol based and conventional superstimulation protocol in buffaloes (Bubalus bubalis).

AIM: To evaluate the follicular dynamics, superovulatory response, and embryo recovery following superstimulatory treatment initiated at estradiol-17ß induced follicular wave emergence and its comparison with conventional superstimulatory protocol in buffaloes. MATERIALS AND METHODS: Six normal cycling pluriparous buffaloes, lactating, 90-180 days post-partum, and weighing between 500 and 660 kg were superstimulated twice with a withdrawal period of 35 days in between two treatments. In superstimulation protocol-1 (estradiol group) buffaloes were administered estradiol-17ß (2 mg, i.m.) and eazibreed controlled internal drug release (CIDR) was inserted intravaginally (day=0) at the random stage of the estrous cycle. On the day 4, buffaloes were superstimulated using follicle stimulating hormone (FSH) 400 mg, divided into 10 tapering doses given at 12 hourly intervals. Prostaglandin F2α analogs (PGF2α) was administered at day 7.5 and day 8, and CIDR was removed with the second PGF2α injection. In superstimulation protocol - 2 (conventional group) buffaloes were superstimulated on the 10(th) day of the estrous cycle with same FSH dose regimen and similar timings for PGF2α injections. In both groups, half of the buffaloes were treated with luteinizing hormone (LH) 25 mg and other half with 100 ug buserelin; gonadotrophin releasing hormone (GnRH) analog at 12 h after the end of FSH treatment. All buffaloes in both protocols were inseminated twice at 12 and 24 h of LH/GnRH treatment. Daily ultrasonography was performed to record the size and number of follicles and superovulatory response. RESULTS: Significantly higher number of small follicles (<8 mm) was present at the time of initiation of superstimulatory treatment in the estradiol group compared to the conventional group (12.5±0.80 vs. 7.3±1.21, respectively, p=0.019), however, the number of ovulatory size follicles (≥8 mm) did not differ significantly between the respective groups (15.5±1.24 vs. 12.2±1.30; p=0.054). Total embryos and transferable embryos recovered were non-significantly higher in the estradiol group compared to the conventional group (5.83±0.86 vs. 4.67±1.16, p=0.328, and 3.67±0.93 vs. 2.67±0.68, p=0.437, respectively). The significant higher proportion of transferable embryos were recovered in buffaloes treated with LH compared to GnRH (73.3% vs. 48.5%; p=0.044). CONCLUSION: The average number of ovulatory size follicles (>8 mm), corpora lutea, and transferable embryos was higher in buffaloes superstimulated at estradiol-induced follicular wave compared to the conventional protocol: Further the percentage of transferable embryos was significantly higher in buffaloes administered with LH compared to GnRH.

Comparative efficacy of E-17ß and GnRH administration on day 0 of a controlled internal drug release (CIDR) based protocol on synchrony of wave emergence, ovulation and conception rates in Murrah buffalos (Bubalus bubalis).

The study was undertaken to compare the efficacy of an estradiol-17ß + CIDR based protocol with the conventional ovsynch + CIDR based protocol for synchrony of wave emergence and ovulation in Murrah buffalos. In group I (n=25), on day 0 (beginning of experiment), buffaloes were administered a CIDR device (1.38 g P4) and concurrently received 1.5 mg E-17ß. On day 9, the CIDR was removed and a prostaglandin (PG) F2α analogue (500 µg) was administered. On day 11, buffaloes were administered a gonadotropin releasing hormone (GnRH) analogue (20 µg) and inseminated twice at 12 h and 24 h following GnRH injections. Group II (n=25) protocol was based on an ovsynch regimen plus CIDR for 7 days followed by double insemination at induced estrus. Group III (n=10) served as control and was not given any hormone on day 0 of the protocol. In groups I, II and III, the duration of new follicular wave emergences were observed on days 4.22 ± 0.12, 3.12 ± 0.33 and 5.14 ± 0.42, respectively. In group I, synchrony of wave emergence was more and the diameter of pre-ovulatory follicles was larger (P<0.05) compared to groups II and III. The first service conception rate (FSCR) was higher (P<0.05) in group I while ovulation rates were not different between groups I and II. In conclusion, more synchrony of wave emergence, larger diameter of dominant follicles and higher first service conception rate was observed following the E-17ß + CIDR based protocol in buffalos.

Synchronization of follicular wave emergence following ultrasound-guided transvaginal follicle ablation or estradiol-17ß administration in water buffalo (Bubalus bubalis).

The aim of this study was to assess the synchrony in follicular wave emergence and subsequent ovulation following dominant follicle ablation or estradiol-17ß administration. Six cycling Murrah buffaloes were sequentially allotted to three groups, that is, control, follicular ablation, and estradiol-17ß groups. For the control group, buffaloes at random stages of estrous cycle were examined daily by transrectal ultrasonography for 14 days and the day of wave emergence was recorded. Following induced luteolysis and ovulation (Day 0), these buffaloes were included in the ablation group. All follicles (>5mm) were ablated on Day 3 or 5 or 7 (n=2 each day). Seven days after the ablation, these buffaloes were administered prostaglandin F2α to induce luteolysis and ovulation. Following this, buffaloes were included in the estradiol treatment group with estradiol administered on similar days as for ablation in the ablation group. Luteolysis was induced nine days after the estradiol injection. All animals of the treatment groups were subjected to transrectal ultrasound and blood samplings daily from treatment to induced ovulation. The follicular waves emerged significantly earlier (P=0.001) in both the ablation (2.1±0.79 days) and estradiol (4.0±0.25 days) treatment groups than the control group (8.3±0.88 days). The deviation from mean day of ovulation was greater (P=0.02) for the control group buffaloes (1.66±0.3 day) than those of the treatment groups (ablation, 0.76±0.2 and estradiol, 0.58±0.2 day). In conclusion, both ablation and estradiol resulted in synchronous emergence of a new follicular wave irrespective of stage at which the treatment was given, with greater synchrony of ovulations in water buffalo.

Correlation of blood flow of the preovulatory follicle to its diameter and endocrine profile in dairy buffalo.

Blood flow of the preovulatory follicle (POF) wall can be used as a predictor of the quality of POF. Our aim was to determine the correlation of blood flow of POF with the POF diameter, and intra-follicular and plasma concentrations of Insulin-like Growth Factor-I (IGF-1) and oestradiol in dairy buffalo. Nine Murrah buffalo subjected to an ovulation synchronization protocol (Ovsynch) were assessed on day 10 of the protocol for diameter and blood flow of POF, followed by the aspiration of follicle fluid. Prior to follicular aspiration, blood samples were obtained from jugular vein for estimation of IGF-1 and oestradiol. The vascularity of POF was determined (Range: 250-967 pixel(2) ) along with intra-follicular and plasma concentration of IGF-1 (Range: 9.3-31.8 ng/ml and 14.7-29.7 ng/ml respectively) and oestradiol (Range: 124.2-447.9 ng/ml and 0.25-1.05 ng/ml respectively). Diameter of the POF was weakly correlated (r = 0.21, p < 0.01) with blood flow to it. As compared to POF diameter, the blood flow of POF had greater positive correlation with intra-follicular and plasma concentrations of hormones (IGF-1 and oestradiol). A strong positive correlation was recorded between intra-follicular IGF-1 and oestradiol. Also, plasma concentrations of oestradiol and progesterone were negatively correlated In brief, assessment of the blood flow of the POF is a non-invasive and reliable indicator of its functional competence as compared to the POF diameter.

Comparison of ovsynch and progesterone-based protocol for induction of synchronized ovulation and conception rate in subestrous buffalo during low-breeding season.

The objective of the present study was to compare the impact of ovsynch and progesterone-based ovulation synchronization protocol on ovarian response and conception in buffalo (n=19) exhibiting subestrus during low-breeding season (maximum ambient temperatures and relative humidity ranging from 36-45°C and 30-80%, respectively). Group I buffalo (n=10) were administered ovsynch protocol (d 0 and d 9, 20 µg Buserelin acetate; d 7, 500 µg Cloprostenol sodium; i.m.) followed by AI on days 9 and 10. During the same period, another group of buffalo (n=9) were administered intravaginal progesterone (1.38 g) for 10 days along with the administration (i.m.) of 500 µg Cloprostenol sodium on day 9 and 20 µg Buserelin acetate on day 11, followed by AI on days 12 and 13. With ovsynch, all the buffalo ovulated in response to 1st GnRH and had functional CL (plasma progesterone, 1.61±0.23 ng/ml; corpus luteum, CL, 11.36±0.67 mm) on day 7. Thereafter, subsequent to 2nd GnRH, five buffalo ovulated within 24 h and the remaining five between 24 to 48 h. In comparison, with progesterone-based protocol, a better synchronization of ovulation (P<0.05) was observed as seven buffalo ovulated between 24 and 48 h and the remaining two between 48 and 72 h following GnRH administration. Moreover, in comparison to ovsynch, conception rate was better with progesterone-based protocol (30 vs. 66.7%; P<0.05). In summary, progesterone-based protocol was superior to ovsynch for synchronization of ovulation and subsequent conception rate in buffalo exhibiting subestrus during the low breeding period.

Improvement of conception rate in postpartum flaxseed supplemented buffalo with Ovsynch+CIDR protocol.

The present study was conducted on lactating Murrah buffalo to assess the effect of crushed flaxseed (a source of omega-3 fatty acids) supplementation (300g/100kg bwt/day for 60 days), over and above the routine feed, on luteolytic signal (PGF2α), luteal function (progesterone) and conception rate. In first experiment, on day 50 post-calving, six non-supplemented buffalo were treated to synchronize time of ovulation using an Ovsynch+Controlled Internal Drug Release (CIDR) protocol followed by intravenous oxytocin treatment (OT; 100IU) on day 15 post-ovulation. Blood samples were collected at 15min interval, 1h before to 4h after OT challenge. Thereafter, the same buffalo were supplemented with flaxseed, treated to synchronize time of ovulation starting on day 35 post-supplementation using the same protocol and subjected to OT treatment and blood sampling on day 15 post-ovulation. The PGF2α response was measured as the venous concentration of 13,14-dihydro-15-keto PGF2α (PGFM). The mean hourly concentration of PGFM subsequent to flaxseed supplemented was less (P<0.05) than in the pre-supplementation period at all the occasions. Flaxseed supplementation did not affect plasma fatty acids and other plasma metabolites except for an increase (P<0.05) in plasma cholesterol and plasma alanine transaminase. In the second experiment, 31 buffalo were randomly assigned to a control (n=16) and flaxseed supplemented (n=15) group. The latter group was supplemented with flaxseed starting from day 15 post-calving. On day 50-post-calving, buffalo of both groups were treated to synchronize time of ovulation among animals as described for the first experiment followed by artificial insemination (AI). Post-AI luteal phase plasma progesterone was greater (P<0.05) in the supplemented group compared to controls. Conception rate on day 63 post-AI was 66.7% in supplemented and 31.2% in controls (P<0.05). The present study indicated the beneficial impact of dietary supplementation of crushed flaxseed on conception rate through attenuation of luteolytic signal and improvement in post-breeding luteal profile.

Induction of ovulation of ovulatory size non-ovulatory follicles and initiation of ovarian cyclicity in summer anoestrous buffalo heifers (Bubalus bubalis) using melatonin implants.

This study was conducted on summer anoestrous buffalo heifers to monitor the efficacy of melatonin for induction of ovulation and ovarian cyclicity. During pre-treatment period of 24 days, the ovarian dynamics of five cycling and 10 summer anoestrous heifers was monitored on each alternate day using a transrectal ultrasound scanner. Thereafter, during treatment period, these 10 anoestrous heifers along with additional seven anoestrous heifers were randomly allocated into non-implanted (n = 5) and implanted (n = 12, one melatonin implant/50 kg, 18 mg melatonin/implant) group. Non-implanted heifers were monitored on each alternate day till the confirmation of second-ovulation in implanted heifers. Pre-treatment period revealed the presence of dominant follicles in anoestrous heifers which attained the diameter comparable with ovulatory follicles of cycling heifers but failed to ovulate and regressed. Between 6 and 36 days (15.3 +/- 2.9 days) post-treatment, all the implanted heifers (p < 0.05) exhibited ovulation of dominant follicles; however none of the non-implanted heifers ovulated during the corresponding period. The first-interovulatory period in implanted heifers ranged between 8 and 28 days (18.0 +/- 1.8 days). The implanted heifers with short (

Estrus induction and fertility rates in response to exogenous hormonal administration in postpartum anestrous and subestrus bovines and buffaloes.

A total of 130 animals (82 cattle, 48 buffaloes) with histories of anestrous 60-90 days post-partum and belonging to different agroclimatic zones of Punjab were subjected to rectal palpation and blood samplings at least three times at weekly intervals. The body condition score (BCS) of each animal was also recorded. The animals were divided into two groups; viz., true anestrous (Gp-I) and subestrus (Gp-II) through rectal palpation of ovaries and plasma progesterone (P4) concentrations. Furthermore, the Gp I and II animals were divided into treatment (Gp Ia, 40 cattle and 16 buffaloes; Gp IIa, 12 cattle and 14 buffaloes) and control groups (Gp Ib, 20 cattle and 8 buffaloes; Gp IIb, 10 cattle and 10 buffaloes). True anestrous animals (Gp Ia) were treated with 3 injections of hydroxyprogesterone caproate (750 mg, i.m.) at 72-hr intervals followed by injection of equine chorionic gonadotropin (eCG; 750 I.U., i.m.) 72 hr after the last progesterone injection. The animals were bred at the first estrus after the induced one. The first service conception rate (FSCR), overall conception rate (OCR), services per conception and pregnancy rate of the true anestrous treated cattle (Gp Ia) were 44.4%, 48.0%, 2.08 and 60.0%, respectively. In the true anestrous control cattle (Gp Ib), only five that were observed to be in estrus failed to conceive. In the anestrous treated buffaloes (Gp Ia), the FSCR, OCR, services per conception and pregnancy rate were 50.0%, 62.5%, 1.6 and 62.5%, respectively. No buffalo amongst true anestrous control (Gp Ib) showed estrus. The subestrus animals (Gp IIa) were administered Prostaglandin F(2alpha) (PGF(2alpha); 25 mg Dinoprost, i.m.) and bred at induced estrus. Amongst the Gp IIa animals, all cattle (100%) and twelve buffaloes (85.7%) responded to treatment. Of these animals, the FSCR and pregnancy rate at induced estrus in the cattle were 50.0% each, whereas they were 66.6% and 57.1%, respectively, in the buffaloes. The subestrus control animals (Gp IIb) remained infertile. In summary, the plasma P(4) profile can be used to differentiate true anestrous and subestrus animals and thus to determine a hormonal therapy. Furthermore, fertile estrus can be induced with hormonal therapy in anestrous and subestrus bovines.