Italian Mediterranean Buffalo Cows Research Articles

Milk quality of Italian Mediterranean buffalo as affected by temperature-humidity index during late spring and summer

A total of 18 Italian Mediterranean buffalo cows were subjected to fortnightly milk sampling from May to July 2022. Air temperature and relative humidity were monitored throughout the trial. Temperature-humidity index (THI) was calculated and ranged from 69 to 79, so that 3 classes were set to study the effect of different THI on milk quality: THI less than 72 (THI<72), THI ranging from 72 to 76 (THI72-76), and THI over 76 (THI>76). Individual milk samples from buffalo cows were analyzed for milk composition and coagulation traits, fatty acid and amino acid composition, and enzymatic and mineral profiles. The THI>76 class registered higher lactose content and poorer milk clot characteristics in comparison with THI<72. Exposure to a THI over 76 resulted in increased SFA and short-chain fatty acid contents in milk, as well as in the highest atherogenic and thrombogenic index values and the lowest essential amino acid content. Moreover, the lowest values of calcium, copper, and selenium contents were found in the milk of buffalo cows exposed to a THI over 72. Results confirm that Italian Mediterranean buffalo express a good level of thermo-tolerance, even though exposure to a daily THI over 76 has a deleterious effect on some nutritional and technological properties of milk.

Evaluation of factors involved in the failure of ovum capture in superovulated buffaloes

The aim of this work was to evaluate factors affecting ovum capture in superovulated buffaloes, by comparing the morphological features of pre-ovulatory follicles and oocytes, the intrafollicular and plasmatic steroid profile, as well as the expression of genes involved in cumulus expansion and steroid cascade in granulosa cells (GCs) and that of genes involved in contraction-relaxation of the oviduct between superovulated and synchronized buffaloes. Italian Mediterranean Buffalo cows were either synchronized by Ovsynch (n = 25) and superovulated (n = 10) with conventional FSH protocol and sacrificed 18 h after last GnRH. Antral follicular count, recovery rate and oocyte quality were recorded, and plasma and follicular fluid were collected for steroid profile determination. In addition, in 10 animals (5/group), GCs were collected to analyse the mRNA expression of gonadotropin receptors (LHR and FSHR) and genes involved in steroid synthesis, as the cytochrome P450 family 19 (CYP19A1) and the steroidogenic acute regulatory protein (STAR). Moreover, oviducts were collected to evaluate the mRNA expression of estrogen receptor 1 (ER1) and the progesterone receptor (PGR), the vascular endothelial growth factor (VEGF) and the VEGF receptors, i.e. the kinase insert domain receptor (FLK1) and the fms related tyrosine kinase 1 (FLT1). No differences were recorded in steroids plasma concentration between synchronized and superovulated animals whereas intrafollicular E2 and P4 concentrations decreased in superovulated group (63.2 ± 10.6 vs 30.3 ± 5.9 ng/mL of E2 and 130.1 ± 19.8 vs 71.6 ± 8.5 ng/mL of P4, respectively in synchronized and superovulated animals; P < 0.05). Interestingly, both the recovery rate (85.7% vs 56.6%, respectively in synchronized and in superovulated animals; P < 0.05) and the percentage of oocytes exhibiting proper cumulus expansion (75% vs 28.1%, respectively in synchronized and in superovulated animals; P < 0.01) decreased in superovulated animals. In addition, the expression of FSHR and CYP19A1 increased while the expression of STAR in GCs decreased (P < 0.05). Finally, in superovulated buffaloes a decreased expression of PGR, ER1, VEGF and its receptor FLK1 in the oviduct was observed. The results suggest that the exogenous FSH treatment impairs steroidogenesis, affecting both the oviduct and the ovarian function, accounting for the failure of ovum capture in superovulated buffaloes.

LH peak and ovulation after two different estrus synchronization treatments in buffalo cows in the daylight-lengthening period

The aim of this study was to determine the timing of ovulation in relation to the LH peak after synchronization using PRID or Ovsynch protocols, to assess the effects of the period of treatment on these parameters and to provide information concerning how to use the two main protocols for fixed-time artificial insemination in buffalo. Forty-eight lactating Italian Mediterranean buffalo cows were used. The buffaloes were treated in various periods as follows: February to March (n = 12 PRID, n = 12 Ovsynch), end of the breeding season, May to June (n = 12 PRID, n = 12 Ovsynch), beginning of low-breeding season according to Italian environmental conditions. To determine the LH, blood samples were taken at 4-hour intervals, starting 24 hours from PRID removal (PRID group) or 12 hours from (PGF2α) injection (Ovsynch group) up to 108 hours. The ovaries were monitored by transrectal ultrasonography to verify ovulation. The LH–ovulation interval was similar in both groups (30.10 ± 1.05 and 32.77 ± 1.15 hours, respectively, in PRID and Ovsynch group). In the PRID group, the timing of ovulation in relation to device removal was 76.83 ± 3.65 hours with a high level of variability among the animals. In the Ovsynch group, we observed a better synchronization of LH peaks and ovulations, and the timing of ovulation in relation to the last GnRH injection was 35.67 ± 1.15 hours. The percentage of animals reaching the LH peak and ovulation was lower (P ≤ 0.05) in May to June (respectively 75.0% and 54.1%) compared to February to March (respectively 95.8% and 83.3%), indicating a reduction of hypothalamus–pituitary responsiveness to the synchronization treatments in the daylight-lengthening period.

Corpus luteum development and function and relationship to pregnancy during the breeding season in the Mediterranean buffalo

The aim of this study was to ascertain corpus luteum (CL) development and function in buffaloes synchronized and mated by artificial insemination (AI) during the breeding season. Italian Mediterranean buffalo cows (n = 43) at 86.5 ± 2.7 days postpartum were synchronized by the Ovsynch-TAI Program and inseminated using frozen thawed semen at 20 and 44 h after the second injection of GnRH. The CL dimensions (diameter and area) and blood flow were examined on Days 5, 10, 15, 20, and 25 after AI by realtime B-mode/colour-Doppler ultrasonography. The resistive index (RI), pulsatility index (PI) and time average medium velocity (TAMV) were recorded at each time, together with CL dimensions. Blood samples were taken on the days of ultrasonography for progesterone (P4) assay by RIA. Data were grouped into pregnant or non-pregnant and retrospectively analyzed by repeated measure ANOVA and correlation analyses. Dimensions of the CL on Days 10, 20, and 25 after AI were greater (P < 0.01) in buffaloes pregnant on Day 45 (n = 18) compared with non-pregnant buffaloes (n = 25). The former buffaloes also showed a greater (P < 0.01) rate of CL growth between Days 5 and 10 after AI. Blood flow to the CL on Day 10 after AI showed a higher TAMV (P < 0.01) and lower RI (P < 0.05) in pregnant buffaloes compared with non-pregnant buffaloes. Negative correlations were observed on Day 10 after AI between CL diameter and RI (r = −0.61; P < 0.01) and PI (r = −0.60; P < 0.01); P4 concentrations and RI (r = −0.46; P < 0.02); and RI and pregnancy (r = 0.45; P < 0.02). Positive correlations were observed between pregnancy and CL size (r = 0.54; P < 0.01), ΔCL diameter between Days 5 and 10 (r = 0.52; P < 0.01), ΔCL area between Days 5 and 10 (r = 0.48; P < 0.015), and ΔP4 between Days 5 and 10 (r = 0.50; P < 0.01). Based on these findings it is concluded that the period between Day 5 and 10 is very important for CL growth and crucial in evaluating pregnancy. Accordingly, the assessment of CL parameters during the period from Day 5 to Day 10 after AI might be used to predict the likelihood of an ongoing pregnancy.

Corpus luteum Function and Pregnancy Outcome in Buffaloes during the Transition Period from Breeding to Non‐Breeding Season

The aim in this study was to investigate corpus luteum function and embryonic loss in buffaloes mated by artificial inseminations (AI) during the transitional period from breeding to non-breeding season. The study was carried out using 288 multiparous Italian Mediterranean Buffalo cows at 110 ± 4 days in milk. The buffaloes were mated by AI after synchronization of ovulation by the Ovsynch-TAI protocol 25 days after AI buffaloes underwent trans-rectal ultrasonography to assess embryonic development. Pregnancy diagnosis was confirmed on Days 45 and 70 after AI by rectal palpation. Buffaloes pregnant on Day 25 but not on Day 45 were considered to have undergone late embryonic mortality (LEM), whilst buffaloes pregnant on Day 45 but not on Day 70 were considered to have undergone foetal mortality (FM). Corpus luteum size and blood flow were determined by real-time B-mode/colour-Doppler on day 10 after AI in 122 buffaloes. The resistive index (RI) and pulsatility index (PI) were recorded at the time. Milk samples were collected on Days 10, 20 and 25 after AI in all inseminated buffaloes for the assay of whey P4 concentrations. Data were analysed by anova. Pregnancy rate on Day 25 after AI was 48.6% (140/288) and declined to 35.4% (102/288) and 30.6% (88/288) by Day 45 and Day 70 respectively. The incidences of LEM and FM were respectively 27.1% (38/140) and 13.7% (14/102). Pregnant buffaloes had greater (p < 0.01) whey concentrations of P4 from Day 20 onwards than buffaloes which showed LEM, whilst P4 in buffaloes that showed FM did not differ from the other two groups on Day 10 and Day 20. Corpus luteum blood flow on Day 10 after AI showed higher RI (p < 0.05) and PI (p = 0.07) values in buffaloes that subsequently were not pregnant on Day 25 compared with pregnant buffaloes. Buffaloes that were not pregnant on Day 45 also had a higher (p = 0.02) RI value on Day 10 than pregnant buffaloes, whilst PI values on Day 10 did not differ for the two groups of buffaloes. It was concluded that blood flow to the corpus luteum on Day 10 after AI influences corpus luteum function as judged by P4 secretion and also embryonic development and attachment in buffaloes.

Influence of replacing corn silage with barley silage in the diets of buffalo cows on milk yield and quality

A 150-day trial was carried out on 40 Italian Mediterranean buffalo cows that, immediately after calving, were equally divided into two homogeneous groups (M and O) based on the number of calving events and previous milk yield. The animals were fed (16 kg dry matter (DM)/head) two isoenergy/isoprotein diets (NEl: 6.39 MJ/kg DM; 15.4 CP% DM), composed of corn (diet M) or barley silage (diet O) concentrate, alfalfa hay, and a vitamin-mineral supplement. The fermentation characteristics of both silage diets were evaluated by an in vitro gas production technique, and their nutritional values were calculated as follows: NEl (MJ/kg DM) = 0.54 + 0.0959 GP + 0.0038 CP + 0.0001733 CP(2), where GP is the gas production after 24 h of incubation (ml/200 mg DM) and CP is the protein content of silage (g/kg DM). The nutritional values of the silages were slightly different (4.16 vs. 4.14 MJ/kg DM for M and O, respectively) likely due to the high content of hemicellulose in the O diet (22.0 vs. 16.9%). Average milk yield did not differ between the groups; instead, milk fat (8.39 vs. 9.06%; P < 0.01) and protein (4.41 vs. 4.60%; P < 0.01) levels were significantly higher in the O group. The results elicit great interest in southern Italy where corn cultivation is adversely affected by the high cost of irrigation.

Follicular dynamics in synchronized Italian Mediterranean buffalo cows

The aim of this study was to evaluate the length and the characteristics of the oestrous cycle in Italian Mediterranean buffalo cows, undergone synchronization of ovulation. The trial was performed on 32 buffaloes synchronized by the Ovsynch Program, which consists of an injection of GnRH on day 0, PGF2α on day 7 and GnRH on day 9. Starting on day 10 (Day 0 of the new cycle). Buffaloes undergone ultrasound examination of the ovaries on alternate days until the following heat. Follicular growth and corpus luteum formation and dimensions were recorded as well as the number of follicular waves. Statistical analysis was performed by ANOVA. Four animals (12.5%) did not show signs of oestrous and were excluded from the trial. The mean length of the oestrous cycle was 23.7±3.4 days. In particular, 1 animal (3.6%) showed an oestrous cycle characterized by 1 follicular wave with a length of 16 days, 17 subjects (60.7%) showed 2 follicular waves with a cycle length of 22.4±2.3 days and 10 buffaloes (35.7%) showed 3 follicular waves with a cycle of 26.8±2.0 days. These results confirm previous reports performed in buffalo species, although the cycle resulted longer in the 3-waves group.

Relationship of body condition score and blood urea and ammonia to pregnancy in Italian Mediterranean buffaloes

The relationship of body condition score (BCS) and blood urea and ammonia to pregnancy outcome was examined in Italian Mediterranean Buffalo cows mated by AI. The study was conducted on 150 buffaloes at 145 +/- 83 days in milk that were fed a diet comprising 14.8% crude protein, 0.9 milk forage units.kg-1 dry matter and a non-structural carbohydrate/crude protein ratio of 2.14. The stage of the oestrous cycle was synchronised by the Ovsynch-TAI programme and blood urea and ammonia levels were assessed on the day of AI. Energy corrected milk (ECM) production and BCS were recorded bi-weekly. The pregnancy risk was 46.7% and was slightly lower in buffaloes with BCS < 6.0 and BCS > 7.5. There were no significant differences in ECM, urea and ammonia between pregnant and non-pregnant buffaloes. However, pregnancy outcome was higher (P = 0.02) in buffaloes with blood urea < 6.83 mmol.L-1. The likelihood of pregnancy for buffaloes with low urea blood level was 2.6 greater than for high urea level and exposure to a high urea level lowered the probability of pregnancy by about 0.25. The findings indicate that buffaloes are similar to cattle and increased blood levels of urea are associated with reduced fertility when animals are mated by AI.

Embryonic mortality in buffaloes synchronized and mated by AI during the seasonal decline in reproductive function

The aim was to determine the factors that contribute to embryonic mortality in buffaloes mated by AI during a period of increasing day length which corresponds to a natural decline in reproductive activity. Italian Mediterranean buffalo cows ( n = 243) showing regular estrous cycles were synchronized using the Ovsynch-TAI program and mated by AI at 16 and 40 h after the second injection of GnRH. Blood samples were collected on Days 10 and 20 after the first AI and assayed for progesterone ( P 4). Pregnancy diagnosis was undertaken on Days 26 and 40 after the first AI using rectal ultrasonography. Buffaloes with a conceptus on Day 26 but not on Day 40 were judged to have undergone embryonic mortality and for these animals uterine fluid was recovered by flushing and analysed for common infectious agents. Estrus synchronization was achieved in 86% of buffaloes and the pregnancy rate on Day 40 was 34%. Embryonic mortality between Days 26 and 40 occurred in 45% of buffaloes and was associated with the presence of significant infectious agents in only 10 buffaloes (8%). Concentrations of P 4 on Day 10 after AI were higher ( P < 0.05) in buffaloes that established a pregnancy than in buffaloes that showed embryonic mortality that was not associated with infectious agents. Similarly, on Day 20 after AI P 4 concentrations were higher ( P < 0.01) in pregnant buffaloes compared with non-pregnant buffaloes and buffaloes that had embryonic mortality. It is concluded that a reduced capacity for P 4 secretion can explain around 50% of embryonic mortalities in buffaloes synchronised and mated by AI during a period of low reproductive activity and that other as yet unidentified factors also have a significant effect on embryonic survival.

138EARLY EMBRYO MORTALITY IN BUFFALO (BUBALUS BUBALIS) DURING THE SPRING PERIOD

Embryo development is linked to several factors, such as intrinsic defects within the embryo, an inadequate maternal environment or failure of maternal response to embryonic signals. Reproductive seasonality in buffalo species (Bubalus bubalis) is responsible for the lower reliability of the pregnancy diagnosis by progesterone assay in spring, probably due to the high incidence of early embryo mortality (EEM) for hormonal reasons (Campanile et al., 1989; Proc. 2th National Meeting ‘Studio dell’efficienza riproduttiva degli animali di interesse zootecnico’: 51–57). The aim of the study was to identify the causes of EEM in buffaloes inseminated during the spring period. Italian Mediterranean Buffalo cows (n=209) in a good state of health were synchronized using the Ovsynch-TAI Program (Pursley et al., 1995 Theriogenology 44, 915–923) and inseminated 16 and 40 hours after the second administration of GnRH. Artificial inseminations were performed by the same technician. Ten to 20 days after the first insemination two blood samples were collected from each animal for assessing P4 levels by RIA. Days 24 and 40 after the first insemination, pregnancy diagnosis was assessed by rectal ultrasonography (Aloka SSD-500) with a 5MHz sector scanner. In animals with EEM a uterine flushing was carried out and recovered fluid was analyzed for the principal infectious agents. Microbiological analyses were carried out according to Quinn et al. (1994 Clinical Veterinary Microbiology, Wolfe Publishing, Mosby, Europe) and a PCR analysis was performed only for Bubaline Herpes Virus (BuHV1), Toxoplasma and Neospora. Statistical analysis was assessed by Student’s t-test. The incidence of cyclic buffaloes after synchronization treatment (94%) and pregnancy rate at 40 days (34.4%) were similar to those reported in a previous study (Neglia et al., 2003 Theriogenology 60, 125–133). EEM between 24 and 40 days was 45% and only in 9 buffaloes (15.3%), these results correlated with the presence of infectious agents (Streptococcus spp. group B and G, Bacillus licheniformis, Proteus mirabilis, Staphylococcus spp.). P4 levels on Day 10 were higher (P&amp;lt;0.05) in pregnant (P) buffaloes than in those with EEM. On Day 20, P4 levels were higher (P&amp;lt;0.01) in P buffaloes than in those with EEM and those not pregnant. It is hypothesized that the reduced activity of hypophysis, correlated with an increase in hours of light, may be responsible for decreased P4 concentrations and EEM in buffalo. Table 1 P4 blood levels in pregnant (P) and non pregnant (NP) buffaloes and in buffaloes with EEM 10 and 20 days after insemination

Comparison of pregnancy rates with two estrus synchronization protocols in Italian Mediterranean Buffalo cows

The aim in this study was to compare two estrus synchronization protocols in buffaloes. Animals were divided into two groups: Group A ( n=111) received 100 μg GnRH on Day 0, 375 μg PGF 2α on Day 7 and 100 μg GnRH on Day 9 (Ovsynch); Group B ( n=117) received an intravaginal drug release device (PRID ®) containing 1.55 g progesterone and a capsule with 10 mg estradiol benzoate for 10 days and were treated with a luteolytic dose of PGF 2α and 1000 IU PMSG at the time of PRID ® withdrawal. Animals were inseminated twice 18 and 42 h after the second injection of GnRH (Group A) and 60 and 84 h after PGF 2α and PMSG injections (Group B). Progesterone (P 4) concentrations in milk samples collected 12 and 2 days before treatments were used to determine cyclic and non-cyclic buffaloes, and milk P 4 concentrations 10 days after Artificial insemination (AI) were used as an index of a functional corpus luteum. Cows were palpated per rectum at 40 and 90 days after AI to determine pregnancies. All previously non-cyclic animals in Group B had elevated P 4 (>120 pg/ml milk whey) on Day 10 after AI. Accordingly, a greater ( P<0.01) relative percentage of animals with elevated P 4 10 days after AI were observed in Group B (93.2%) than in Group A (81.1%). However, there was no difference in overall pregnancy rates between the two estrus synchronization protocols (Group A, 36.0%; Group B 28.2%). When only animals with elevated P 4 on Day 10 after AI were considered, pregnancy rate was higher ( P<0.05) for animals in Group A (44.4%) than Group B (30.3%). The findings indicated that treatment with PRID ® can induce ovulation in non-cyclic buffalo cows. However, synchronization of estrus with Ovsynch resulted in a higher pregnancy rate compared with synchronization with PRID ®, particularly in cyclic buffalo.

Repeated ovum pick-up in Italian Mediterranean buffalo cows

The potential of the ovum pick-up technique, used over a long period, was evaluated in 6 Italian Mediterranean buffalo cows that had more than 500 d open. The cows were submitted to ovum pick-up twice weekly for 2 mo. An additional 2-mo cycle of ovum pick-up was performed in 3 of the buffalo. The ovum pick-up sampling did not affect the resumption of reproductive activity of these animals. In fact, all the buffalo conceived, on average, 47.5 ± 27.5 d after the last ovum pick-up. An average of 5.48 follicles was punctured, and 2.71 oocytes were collected per session. However, only 53.5% of these oocytes were suitable for in vitro embryo production. The number of punctured follicles differed between individual cows. There were no differences in the number of collected oocytes or in the recovery rates. The number of punctured follicles, the number of collected oocytes and the recovery rate were similar in the first and second months; the quality of the oocytes was, however, better in the second than in the first month (P < 0.05). The increasing interval between 2 consecutive ovum pick-up sampling (intersession interval) caused an increase of the percentage of large follicles. Moreover, the increase of the intersession interval from 4 to 5 d decreased the quality of the collected oocytes (P < 0.05). The efficiency of in vitro production of embryos to expanded blastocysts was 16.7%.