Pregnancy In Buffaloes Research Articles

Endometrial transcript profile of progesterone-regulated genes during early pregnancy of Water Buffalo (Bubalus bubalis).

Progesterone (P4 ) plays a key role in the establishment and maintenance of pregnancy in most mammals. Unravelling the expression of progesterone-regulated genes can expand the understanding of the embryonic mortality. Accordingly, we studied the relative mRNA expression of the P4 -regulated genes in the buffalo. Uteri were collected from the abattoir and categorized into nonpregnant late luteal phase, stage I (28-38th days of gestation) and stage II (48-56th days of gestation) of pregnancy (n=6/group). After extraction of total RNA from the endometrial tissues, we carried out qRT-PCR for determining the relative mRNA expression of the P4 -regulated genes using nonpregnant late luteal phase as calibrator group. The expression of LGALS3BP (essential for maternal recognition of pregnancy) gene was found to be significantly upregulated (p<0.05), while MUC1 (important for embryo attachment) gene was downregulated in stage I and II of pregnancy. We observed no significant change in the expression of LGALS1, LGALS9 and CTSL genes. The SLC5A11 and SLC2A1 genes (involved in the transport of glucose to endometrium) in early pregnancy were upregulated in the pregnancy stage I (p<0.05) relative to nonpregnant late luteal phase. The CST3 gene was significantly upregulated in pregnancy stage II (p<0.01). These results provide molecular insights into the specific pathways involved in foeto-maternal communication during early pregnancy in buffaloes.

Recombinant interferon stimulated protein 15 (rISG15) as a molecular marker for detection of early pregnancy in Bubalus bubalis

Early and accurate diagnosis of pregnancy in animals is important for improving the reproductive management of livestock. The buffalo (Bubalus bubalis) is the most important dairy animal in India, but there are reproductive problems resulting from extended calving interval and ovulation occurring in the absence of behavioral estrus. The lack of simple methods for early pregnancy diagnosis intensifies these problems. The present study, therefore, was conducted to ascertain the role of the interferon-stimulated gene, (ISG), 15 in pregnancy detection. The anti-ISG15 Mab based ELISA was developed that could be used for detecting pregnancy at 18 to 20 days after artificial insemination (AI). The ISG15 protein was isolated from a pregnant buffalo and was amplified, and cloned in Escherichia coli by using coding region primers. The ISG15 gene was expressed in the host Escherichia coli BL21 (DE3), and the protocol was standardized for optimum gene expression. Using immortal hybridoma (fused myeloma and B cells) cells, a highly specific and sensitive antibody, anti-ISG15 mAb, for detecting ISG15 (protein) in the serum of pregnant buffaloes was obtained. A blocking ELISA was developed using the anti-ISG15 mAb to detect pregnancy in buffalo within 18 to 21 days after AI. The ISG15 gene was upregulated (P < 0.05) in pregnant buffalo at 18 to 21 days of pregnancy. This assay has an overall diagnostic accuracy of 75.0%. It, therefore, is concluded that recombinant ISG15 retains the potential for detecting pregnancy in B. bubalis and may have applications in ELISA kits for pregnancy detection in closely related species.

Identification of microRNAs in corpus luteum of pregnancy in buffalo (Bubalus bubalis) by deep sequencing.

This study was aimed to identify miRNAs of corpus luteum (CL) in buffaloes during pregnancy. For this study, CL (n=2) were collected from gravid uteri of buffalo and RNA was isolated. Following this, the purity and integrity of RNA was checked and used for deep sequencing using Illumina Hiseq 2500 platform. The reads' quality was checked prior to in silico analyses viz. identification of conserved, novel and target of miRNAs. In this study, out of identified miRNAs (3018), 3013 were known and 5 were novel miRNAs on alignment with reference genomes. In addition, prediction of putative target genes for identified abundant miRNAs revealed several genes viz. HOX, KLF4, NCOR2, CDKN2Z, MAPK7, COX2, PPARA, PTEN, ASS3A, ELK1, CASP3, BCL211, MCL1, CCND2, Cyclin A2 and CDC25A during early pregnancy in buffalo. These predicted target genes have been associated with various cellular house-keeping processes including apoptosis. In conclusion, this study reports the identification of conserved and novel microRNAs (miRNAs) in CL during pregnancy in buffalo by deep sequencing.

Predicting early pregnancy in Egyptian buffalo cows via measuring uterine and luteal blood flows, and serum and saliva progesterone.

Various methods are being employed to detect early pregnancy in domestic animals. This study aimed to predict early pregnancy in buffaloes via measuring the corpus luteum (CL) diameter, the luteal blood flow (LBF) area, the uterine blood flow (UBF) vascularization area, and progesterones in saliva and serum for non-pregnant (NPBs, N=12) and pregnant (PBs, N=12) buffaloes. The results revealed that the CL diameter and the luteal color blood flow blue and red (P=0.0001) areas of the pregnant animals kept increasing from day 1 to day 35 of the gestation period, but it decreased in NPBs on day 21 after reaching a peak from ovulation to day 18. Interestingly, the UBF of the pregnant buffaloes (PBs) kept increasing (P=0.0001) from ovulation to day 42. The difference of the CL diameter (P=0.03) and the LBF color blue vascularization area (P=0.002) between PBs and NPBs became clear from day 14 after ovulation, though the difference of UBF between PBs and NPBs became markedly obvious from day 7 after breeding. Both saliva (P=0.001) and serum (P=0.0001) progesterones of PBs continued increasing (P=0.0001) from day 14 to day 35, but those of NPBs started decreasing (P=0.0001) from day 14 and reached low values on day 21. Therefore, measuring saliva progesterone in addition to the high LBF (day 14) and UBF (day 7) of the pregnant buffaloes using a Doppler ultrasound could be applicable as noninvasive methods to detect early pregnancy and to improve reproductive management of buffaloes.

Repertoire of noncoding RNAs in corpus luteum of early pregnancy in buffalo (Bubalus bubalis).

Aim:The present study was designed to identify other noncoding RNAs (ncRNAs) in the corpus luteum (CL) during early pregnancy in buffalo.Materials and Methods:For this study, CL (n=2) from two buffalo gravid uteri, obtained from the slaughter house, was transported to laboratory after snap freezing in liquid nitrogen (−196°C). The stage of pregnancy was determined by measuring the crown-rump region of the fetus. This was followed by isolation of RNA and deep sequencing. Post-deep sequencing, the obtained reads were checked and aligned against various ncRNA databases (GtRNA, RFAM, and deep guide). Various parameters, namely, frequency of specific ncRNAs, length, mismatch, and genomic location target in several model species were deciphered.Results:Frequency of piwi-interacting RNAs (piwi-RNAs), having target location in rodents and human genomes, were significantly higher compared to other piwi-RNAs and ncRNAs. Ribosomal RNAs (rRNAs) deduced had nucleotides (nts) ranging from 17 to 50 nts, but the occurrence of small length rRNAs was more than lengthier fragments. The target on 16S rRNA species confirms the conservation of 16S rRNA across species. With respect to transfer RNA (tRNA), the abundantly occurring tRNAs were unique with no duplication. Small nucleolar RNAs (snoRNAs), identified in this study, showed a strong tendency for coding box C/D snoRNAs in comparison to H/ACA snoRNAs. Regulatory and evolutionary implications of these identified ncRNAs are yet to be delineated in many species, including buffaloes.Conclusion:This is the first report of identification of other ncRNAs in CL of early pregnancy in buffalo.

Electrochemical detection of estrus specific phenolic compound p-cresol to assess the reproductive phase of certain farm animals

p-Cresol (4-Methylphenol) is one of the naturally occurring volatile phenolic compounds in the female mammalian animals urine during estrus, the period of time when the animals become sexually receptive and release specific odors that are induced by reproductive hormones. We have developed an electrochemical sensor for p-cresol detection using hydroxyapatite nanoparticles (HA NP) for the first time. The HA nanoparticles have been synthesized by microwave irradiation method using plectranthus amboincus plant-extract as solvent and its properties have been characterized by XRD, FTIR, and SEM methods The plant-extract-mediated HA NP modified glassy carbon electrode (GCE) exhibited improved electrocatalytic behavior towards p-cresol sensing compared to bare GCE and pristine HA NP/GCEs. Under optimized conditions, the proposed sensing platform permits the detection of p-cresol over a wide range of 0.275–23.5×10−6M with the lowest detection limit of 116×10−9M (S/N=3) in phosphate buffer solution (pH 6.0).The fabricated sensor has high selectivity towards p-cresol in the presence of potential interferents such as nitrophenols, bisphenol A, catechol, hydroquinone, uric acid, glucose, folic acid, ascorbic acid and metal ions Na+, K+ and Mg2+. The sensor has been successfully used to detect p-cresol in female buffalo urine which eventually indicates its readiness for artificial insemination. This device will be helpful to enhance the success rate of pregnancy in buffalo and other farm animals.

Expression and molecular cloning of interferon stimulated genes in buffalo (Bubalus bubalis)

Buffalo, the most important livestock species in tropical India, remains to be a poor breeder mainly due to embryonic mortality (65%) occurring mostly between 16 and 18 days of pregnancy. Early and accurate diagnosis of pregnancy can thus become a boon for successful herd management in buffalo. However, most of the currently available methods allow diagnosis only after 30 days post AI. Interferon tau (IFNT), the first pregnancy recognition signal in ruminants is one such molecule, which stimulates expression of various Interferon stimulated genes (ISGs) in the peripheral blood mononuclear cells (PBMC's) concomitant with IFNT signaling which occurs around maternal recognition of pregnancy (MRP). Hence, the study was planned to demonstrate the expression dynamics of ISGs (OAS1, MX1, MX2 and ISG15) in PBMCs during peri-implantation period in buffalo and also molecular cloning and expression of suitable ISG coded protein (s) in suitable host. Blood was collected from two groups of multiparous buffaloes: Group1: (n = 10) inseminated/pregnant (Experimental) and Group2: (n = 10) anestrous/non pregnant (Control). The expression profile of ISGs was then analyzed using real time qPCR. Expression profile of most ISGs was observed to increase through day 14 to day 20 post AI and declined thereafter. On the basis of differential gene expression at day 18 post AI, OAS1 and MX2 were identified as suitable ISG candidate biomarkers for accurate pregnancy diagnosis within 18 days post AI. Molecular cloning and expression of selected ISGs in a suitable prokaryotic expression vector was done thereafter. Bulk expression of the recombinant proteins was done and purified by affinity chromatography and confirmed by Western blot using Mouse Monoclonal His-probe antibodies. To conclude, as OAS1 and MX2, showed distinct differential expression at day 18 post AI, they may serve as ideal biomarkers for detection of early pregnancy in buffalo.

Pregnancy-associated glycoproteins (PAGs) concentrations in water buffaloes (Bubalus bubalis) during gestation and the postpartum period

For the first time in literature this study describes the pregnancy-associated glycoprotein (PAG) profile of buffalo cows during gestation and the post-partum period using antiserum raised against PAG-molecules purified from buffalo placenta (AS#860).Ninety-eight buffalo cows, belonging to a buffalo herd subjected to a synchronization and artificial insemination (AI) program, were enrolled in this study. Blood samples were taken on days 0 (AI), 23, 25, 28, 30 and then biweekly until the end of pregnancy. Pregnancy was confirmed by ultrasonography on days 28 and 45, and by rectal palpation from day 60 onwards. Blood samples were suspended for the non-pregnant cows on day 45, while the blood of 20 buffaloes that had calved was tested every five days from the day of calving until day 50 post-calving. A cut-off value of 1.0 ng/mL was used in order to discriminate between pregnant and non-pregnant buffaloes. We used Linear Mixed models after Log(x+1) transformation to analyse the PAG concentrations.Fifty-two buffalo cows had become pregnant out of 98 synchronized (53%) and 46 remained non-pregnant (47%) as shown by ultrasonography and the PAG analysis. Significant differences (P < 0.001) in PAG concentrations were observed between the pregnant and non-pregnant buffaloes from day 23 as the PAG of the non-pregnant cows was always close to zero. Conversely, the PAG of the pregnant cows increased progressively from day AI until day 105 post-insemination and then stabilized until the end of pregnancy. Regarding pregnancy diagnosis, the sensitivity of PAG-RIA 860 system (ability of the test to correctly identify pregnant buffalo) ranged from 23% on day 23–98% on day 28 post AI; the specificity (ability to correctly identify non-pregnant buffaloes) was 100% throughout the sampling period. PAG progressively decreased from parturition to day 25 post-partum; from day 30 post-partum, the concentrations fell below 1 ng/mL and were close to 0 on the last day of observation (50 d post-partum).In conclusion, our results showed that RIA-860 is highly accurate for diagnosing pregnancy in buffaloes starting from day 28 of gestation. Furthermore, the rapid disappearance of PAG concentration after calving means that a cut-off limit in post-partum for detecting a new pregnancy is not required.

Identification of serum protein markers for early diagnosis of pregnancy in buffalo.

Improper or delayed pregnancy diagnosis has significant impact over animal production, particularly in buffaloes which inherently suffer from several reproductive inefficiencies. Thus the present study has undertaken to identify serum protein markers pertaining to early pregnancy diagnosis in buffaloes. Serum samples were collected from 10 pregnant Murrah Buffalo heifers at weekly intervals from days 0-35 post-artificial insemination and from 12 inseminated non-pregnant cyclic buffalo heifers on days 0, 7, 14 and 21. Two-dimensional gel electrophoresis and densitometric analysis revealed the presence of five protein spots showing average density fold change of ≥4 during early pregnancy. Mass spectrometry analysis identified these up-regulated proteins as anti-testosterone antibody light chain, apolipoprotein A-II precursor, serum amyloid A, cytokeratin type II, component IV isoform 1, which are have established roles in embryogenesis, but over-expression of the fifth identified protein immunoglobulin lambda light chain in pregnancy has been elucidated as a novel finding in the current study. Further, with bioinformatics analysis, potential antigenic B-cell epitopes were predicted for all these five proteins. An antibody cocktail-based approach involving antibodies against all these five up-regulated entire proteins or their epitopes could be developed for early detection of pregnancy in buffaloes. © 2016 Japanese Society of Animal Science.

Serum MX2 Protein as Candidate Biomarker for Early Pregnancy Diagnosis in Buffalo.

Interferon-tau (IFN-τ)-induced molecular markers such as ubiquitin-like modifier (ISG15), 2',5'-oligoadenylate synthetase 1 (OAS1) and myxovirus resistance genes (MX1 and MX2) have generated immense attention towards developing diagnostic tools for early diagnosis of pregnancy in bovine. These molecules are expressed at transcriptional level in peripheral nucleated cells. However, their presence in the serum is still a question mark. This study reports sequential changes in expression of MX2 transcript in whole blood and serum MX2 protein level on days 0, 7, 14, 21, 28 and 35 in pregnant (n=9) buffalo heifers, and on days 0, 7 and 14 in non-inseminated (n=8) and inseminated non-pregnant (n=10) control animals. In non-inseminated and inseminated non-pregnant heifers, the differential expression of MX2 transcript and MX2 protein level remained similar between day 7 and 14 post-oestrus. However, in pregnant heifers, on 14th and 28th day post-insemination MX2 transcript was 16.38±1.57 and 28.16±1.91 times upregulated as compared to day 0. Similarly, serum MX2 protein concentration followed analogous trend as MX2 transcript and increased gradually with the progression of pregnancy. Correlation analysis between expression of MX2 transcript and its serum protein level showed a significant positive correlation in pregnant animals, while it was random in other two groups. Therefore, MX2 surge at transcriptional and serum protein level after day 14-28 of pregnancy in buffalo holds potential for its use in early pregnancy detection.

Effect of timing of insemination after CIDR removal with or without GnRH on pregnancy rates in Nili-Ravi buffalo

There are limited studies available on when to inseminate, if using CIDR or CIDR-GnRH protocols for optimal fertility in buffalo. Therefore, the objective of the present study was to determine the optimum time of AI in relation to CIDR removal with or without GnRH in buffalo. All buffalo (n=201) received CIDR on Day 0, PGF2α on Day 6 and CIDR were removed on Day 7. In 50 of these buffalo GnRH was administered 36h after CIDR removal. Furthermore, CIDR (n=151) and CIDR-GnRH (n=50) were randomly divided to receive timed artificial insemination (TAI), either at 48, 60 or 72h after CIDR removal. Ultrasonography was performed for follicular development and for pregnancy diagnosis. The results revealed that, mean interval to ovulation was shorter (P<0.05) in buffalo receiving CIDR-GnRH than CIDR (68.40±1.73 compared with 76.13±1.66h, respectively). The pregnancy rates were higher (P<0.05) in buffalo inseminated either at 48 (50%) or 60 (59%) than at 72h (18%) in CIDR-GnRH protocol; whereas, in CIDR buffalo pregnancy rates were higher (P<0.05), at 60 (37%) or 72 (40%) than at 48h (10%). In conclusion, the optimal time of AI is between 48–60h in CIDR-GnRH and between 60–72h in CIDR protocol for enhancing fertility in buffalo.

Ovarian follicular development, estradiol and progesterone profile during different days of early pregnancy in buffaloes *

Improvement of reproductive performance of buffaloes requires a better understanding of the mechanisms controlling ovarian follicular growth and development. To study the follicular dynamics and ovarian steroid profile during early pregnancy, 6 buffaloes were selected on the basis of non return rate after insemination and subjected for ultrasonography to confirm the pregnancy as well as follicular development. The mean number of medium, large and total follicles was significantly lower on days 30, 40, 50 and 60 than day 20 of pregnancy indicating reduction in the follicular population with the advancement of pregnancy. There was no significant difference in the diameter of small and medium follicles on different days of pregnancy, however, large follicle significantly decreased with advancement of pregnancy. The progesterone levels fluctuated between 2.52±0.26 and 3.12±0.44 ng/ml until third month of pregnancy. The mean serum estradiol was significantly higher on day 0 and reduced into basal level on first and second month and remained low throughout the study period. In conclusion, on day 20 to 60 of pregnancy, the follicular development is continuous and the number and diameter of larger follicle is decreased as pregnancy advances in buffaloes.

Changes in sera proteome in relation to day of pregnancy in early pregnant buffaloes

The present study was planned for studying the sequential changes in early pregnant buffalo for the development of diagnostics for early pregnancy. In the present study, sera samples obtained at weekly intervals from early pregnant (day 0 to day 42 post-AI) buffaloes, and on days 0, 7 and 14 from non-pregnant cyclic buffaloes, were subjected to depletion of high abundant proteins followed by 2-dimensional gel electrophoresis and densitometric analysis. At least 65 2-D gel spots exhibited up-regulation, down-regulation or specific appearance at a specific stage during early buffalo pregnancy, except for the spots correlating with the high abundant proteins’ location. Comparison with ExPASy and NCBI databases matched 48 of these spots with known proteins, but with varying degrees of confidence in terms of Mascot score and the species. Although high abundant proteins were depleted before 2-D electrophoresis, yet in some of the picked spots isoforms of common abundant proteins, viz. serum albumin, igg, serrotransferrin, complement and mhc molecules, were found. synaptojanin-1, apolipoprotein a-1, apolipoprotein b, keratin 10 and von Willebrand factors were some of the proteins identified in these spots, which have a documented role in embryogenesis and early pregnancy.

Bioinformatics and functional analysis of proteins in serum of early pregnant buffaloes

Bioinformatics using different web based resources was done to classify 2–D gel resolved novel/ up- or down- regulated proteins (n=48) detected in serum of early pregnant buffaloes and find out interactions among them. Proteins were searched for their gene symbols on the ExPASy and the NCBI web portals and then PANTHER software was used to classify them. These proteins were primarily involved in regulatory, catalytic, immune regulation, cell differentiation and transporter functions. Among molecular functions, majority of proteins was involved in either catalytic or binding activities (29.5% each); receptor activity (13.6%), transporter activity (9.1%), enzyme regulator (6.8%) and structural molecule activity (6.8%) besides ion-channel and antioxidant activities. Based on biological functions, maximum (17.6%) proteins were found involved in metabolic function, followed by cellular processes (15.7%). Almost 10.8% proteins were involved in cell communications and 9.8% in transport functions, the rest were involved in immune and developmental processes. To these proteins 23 pathways were assigned, most predominantly Wnt signalling, inflammation - mediated by chemokine and cytokine signalling, and blood coagulation pathways. Other proteins were involved in as many as 20 pathways. A single layer extension to make connections for preparing protein-protein interaction map and network analysis of these bovine proteins, revealed that 9 of these proteins were interacting with each other in 6 different ways - interactions affecting molecular transportation, level of expression, regulation of expression, binding of proteins, interactions resulting in direct regulation and direct regulation inhibition and those leading to modification of the proteins. These findings suggested that the proteins identified with up-regulation, down-regulation or specific appearance at a particular stage during early buffalo pregnancy, primarily act interdependently in the functions involving cellular growth and differentiation.

In silicoprediction of buffalo (Bubalus bubalis) pregnancy associated glycoprotein-1 signaling pathway

The present study was conducted to predict and analyze signaling pathway of buffalo pregnancy associated glycoprotein-1 in silico. Various databases viz. String database, Panther pathways, Biocarta pathways were used for deducing signaling of buffalo pregnancy associated glycoprotein-1. Analysis reveals buffalo pregnancy associated glycoprotein-1 exerts their biological function by interacting with cytokines viz. placenta growth factor and SP1 transcription factor. Placental growth factor belongs to the vascular endothelial growth factor superfamily. Vascular endothelial growth factor is mainly involved in cellular proliferation, migration, differentiation, angiogenesis acting through protein kinase-C signaling pathway. SP1 transcription factor is mostly involved in mediators of transcription and signal transduction during cellular process acting through the SMAD factors. In toto pregnancy associated glycoprotein-1 through these factors may exert its biological functions of angiogenesis, endothelial cell growth, proliferation, migration and differentiation enhancing embryonic growth and development. In conclusion from this study the signaling pathway of buffalo pregnancy associated glycoprotein-1 (PAG-1) was predicted in silico.

THE PRENATAL DEVELOPMENT IN SWAMP BUFFALO (BUBALUS CARABANENSIS)

Summary There are morphological and reproductive physiological differences between swamp buffalo (Bubalus carabanensis) and river buffalo (Bubalus bubalis). The development of fetus weight and fetus biometry was reported in river buffalo and other animals but not in swamp buffalo. The aim of this study was to describe the inherent variability in fetus related measurements during swamp buffalo pregnancy. The data is based on measurements of 267 fetuses and 5 new born claves from swamp buffalo. The results show that a significant linear correlation exists between estimated age of fetuses and parameters of fetus sizes. There were correlations between crown-rump length (CRL) and other fetal parameters, as well as between fetus weight and its parameters. In conclusion, our data indicated that the feasibility and value of fetal measures in swamp buffaloes being used for the evaluation of fetal development.

Interaction analysis of buffalo pregnancy associated glycoprotein-1 in silico

Pregnancy associated glycoprotein-1, a member of the aspartic proteinase family possesses placentogenic and embryo protective functions in numerous domestic species. The present study was conducted to deduce binding and interaction properties of buffalo pregnancy associated glycoprotein-1 (PAG-1) in silico. Buffalo PAG-1 DNA, RNA binding sites deduced using BindN server revealed buffalo PAG-1 protein sequence possess 93 residues with 80% and 56.96% DNA binding specificity and sensitivity, respectively. RNA binding sites are also restricted to identical clusters of amino acid residues as DNA binding sites with 78 residues with RNA binding potential with 80% specificity and 53.95% sensitivity. The potential domains with a high degree of DNA and RNA binding property are present at conserved microsequences residues of buffalo PAG-1 protein sequence. Ligand binding properties from PDBSUM database reveal buffalo PAG-1 possesses ten clefts having potential ligand binding sites consisting of aliphatic and positive amino acid residues. Analysis with the STRING database showed buffalo PAG-1 interaction with cytokines viz. phosphoprotein associated with glycosphingolipid microdomains 1, placenta growth factor precursor, alpha-fetoprotein precursor and SP1 transcription factor. These factors are found to be active during the embryonic stage exerting their functions through angiogenesis, endothelial cell growth, proliferation, migration and differentiation. In conclusion this study reports the various binding properties and putative functional interactions of buffalo pregnancy associated glycoprotein-1 with other cytokines for exerting its biological action.

Uterine blood flow during various stages of pregnancy in dairy buffaloes using transrectal Doppler ultrasonography

Reference data to assess function and perfusion of uteroplacental tissues by assessment of uterine blood flow parameters is not available in buffaloes. The objective of the present study was to assess and quantify blood flow parameters at various stages of gestation. A total of 54 Murrah buffaloes, six animals at every month of gestation, from two months to ten months, were subjected to transrectal Doppler ultrasonography to assess uterine blood flow in both the middle uterine arteries. Resistance and pulsatility indices, velocity and volume of blood flow, diameter of the artery and notch signal were measured for different waveforms. The resistance index value was positively correlated with pulsatility index (r=0.94, P<0.05) but negatively correlated with all other parameters for both arteries, ipsilateral and contralateral to the fetus. With a significant advancement of pregnancy, there was reduction in resistance index with a significant increase in velocity, volume of blood flow and diameter in both arteries. There was a significant increase in the volume and velocity of blood flow (P<0.05) during the last trimester with increased growth demands of the fetus. The notch signal disappeared by 20–24 weeks. Thus, transrectal Doppler ultrasongraphy proved to be a useful non-invasive method to assess the uterine blood flow during pregnancy in buffaloes. This could be a valuable tool to ascertain hemodynamic changes in complicated pregnancy (e.g. abnormalities in uterine blood flow/placenta/fetus) and its extent of deviation from normal.

Studies on physical and rheological properties of cervico-vaginal mucus during early pregnancy in buffaloes (Bubalus bubalis)

Aim: The present work was carried out to study and compare the variation in physical and rheological (study of stretch and elasticity) properties of cervico-vaginal mucus of pregnant and non pregnant animals which could be used for early pregnancy diagnosis in buffaloes. Materials and Methods: Cervico-vaginal mucus samples were obtained from 30 buffaloes (n=12 pregnant, n=18 non pregnant) and a comparison was made between values of different parameters obtained from pregnant and non pregnant animals. Transparency, consistency, and elasticity of mucus were studied during early pregnancy. Results: Transparency and consistency did not show any remarkable difference between pregnant and non pregnant animals during any phase of estrous cycle. However, rheological property i.e. elasticity, showed a significant difference in its value on th th 9 day of estrous cycle. Spinnbarkeit value decreased significantly in pregnant animals on 9 day (3.9±0.45 cm), whereas non th pregnant group did not show a remarkable difference (8.4±0.43). Spinnbarkeit value was unidentifiable from 12 day onward in both the groups. Conclusion: These results conclude that significant variation in mucus elasticity could be helpful in designing user friendly tool for early pregnancy diagnosis in buffalo in future.

Equine chorionic gonadotropin improves the efficacy of a timed artificial insemination protocol in buffalo during the nonbreeding season

Two experiments were conducted to evaluate the effects of equine chorionic gonadotropin (eCG) treatment on ovarian follicular response, luteal function, and pregnancy in buffaloes subjected to a timed artificial insemination (TAI) protocol during the nonbreeding season. In experiment 1, 59 buffalo cows were randomly assigned to two groups (with and without eCG). On the first day of the synchronization protocol (Day 0), cows received an intravaginal progesterone (P4) device plus 2.0 mg estradiol benzoate im. On Day 9, the P4 device was removed, all cows were given 0.150 mg PGF2α im, and half were given 400 IU eCG im. On Day 11, all cows were given 10 μg of buserelin acetate im (GnRH). Transrectal ultrasonography of the ovaries was performed on Days 0 and 9 to determine the presence and diameter of the largest follicle; between Days 11 and 14 (12 hours apart), to evaluate the dominant follicle diameter and the interval from device removal to ovulation; and on Days 16, 20, and 24 to measure CL diameter. Blood samples were collected on Days 16, 20, and 24 to measure serum P4. In experiment 2, 256 buffaloes were assigned to the same treatments described in experiment 1, and TAI was performed 16 hours after GnRH treatment. Pregnancy diagnosis was performed by ultrasonography 30 days after TAI. Treatment with eCG increased the maximum diameter of dominant follicles (P = 0.09), ovulation rate (P = 0.05), CL diameter (P = 0.03), and P4 concentrations (P = 0.01) 4 days after TAI, and pregnancy per AI (52.7%, 68/129 vs. 39.4%, 50/127; P = 0.03). Therefore, eCG improved ovarian follicular response, luteal function during the subsequent diestrus, and fertility for buffalo subjected to a TAI synchronization protocol during the nonbreeding season.