Search

The present study was planned to find out the magnitude of environmental factors affecting udder type traits in Sahiwal cattle. Fourteen type of udder traits were measured for each of randomly selected 100 lactating cattle maintained at Livestock Research farm, NDRI, Karnal, India. The data were analysed by least squares methods using the general linear model procedures including parity, stage of lactation and season of calving at classification as fixed effects. Traits viz. Udder balance, Rear Udder Height, Teat Diameter, Teat Length, Central Ligament, Udder Circumference, Udder Width and Teat Circumference were significantly (p

A lactation curve can reveal abundant information about the pattern and persistency of lactation in an individual dairy animal. Identifying buffalo with particular pattern and persistency of lactation milk yield can serve as additional selection criterion for a dairy herd. Wood’s incomplete gamma function fitted well to the monthly test-day milk, fat% and SNF% yields of Murrah buffaloes and can be used in prediction of milk and milk constituents yield with higher accuracy. We also report that exponential function (Wilminks model) also can be used in prediction of milk constituents in Murrah buffaloes when constant k is changed in the model from 0.05 to 0.06 or 0.07. Thus, these functions can be used to make breeding and management related decisions particularly selection of desired and persistent buffaloes and thus improving the production of the herd by culling non-desired type of Murrah buffaloes.

Murrah breed of buffalo is an excellent dairy germplasm known for its superior milk quality in terms of milk fat and solids-not-fat (SNF); however, it is often reported that Indian buffaloes had lower lactation and fertility potential compared to the non-native cattle of the country. Recent techniques, particularly the genome-wide association studies (GWAS), to identify genomic variations associated with lactation and fertility traits offer prospects for systematic improvement of buffalo. DNA samples were sequenced using the double-digestion restriction-associated DNA (RAD) tag genotyping-by-sequencing. The bioinformatics pipeline was standardized to call the variants, and single-nucleotide polymorphisms (SNPs) qualifying the stringent quality check measures were retained for GWAS. Over 38,000 SNPs were used to perform GWAS on the first two principal components of test-day records of milk yields, fat percentages, and SNF percentages, separately. GWAS was also performed on 305 days’ milk yield; lactation persistency was estimated through the rate of decline after attaining the peak yield method, along with three other standard methods; and breeding efficiency, post-partum breeding interval, and age at sexual maturity were considered fertility traits. Significant association of SNPs was observed for the first principal component, explaining the maximum proportion of variation in milk yield. Furthermore, some potential genomic regions were identified to have a potential role in regulating milk yield and fertility in Murrah. Identification of such genomic regions shall help in carrying out an early selection of high-yielding persistent Murrah buffaloes and, in the long run, would be helpful in shaping their future genetic improvement programs.

Four divergent rations namely low protein-low energy (LP-LE), low protein-high energy (LP-HE), high proteinlow energy (HP-LE) and high protein-high energy (HP-HE) were formulated to supply two levels of digestible crude protein (DCP) (100 and 75%) and 2 levels of energy (TDN) (100 and 75%) for twenty four growing yaks (12 male and 12 female) of uniform age and body weights randomly divided into 4 groups of 6 animals (3 male and 3 female) in each. At random one dietary treatment was allotted to each group for a period of one year. At the end of feeding trial, average daily gain and feed conversion efficiency were highest in HE-HP group followed by HP-LE, LP-HE and LP-LE groups though their differences were statistically non-significant. The blood glucose level was highest in HP-HE group in comparison to the other groups; but the trends of plasma protein, albumin and urea levels were similar among the groups. The study also revealed that high protein-high energy ration is the suitable dietary combination for growing yak calves to support an average body weight gain of 371.8 g/day during 250 to 300 kg of body weight and could advance their sexual maturity.

The present study highlighted the state of climate change induced social vulnerability of the districts of Arunachal Pradesh. For the purpose of assessment of one of the most fragile ecosystems of the eastern Himalaya, the ‘Integrated Vulnerability Assessment Approach’ and IPCC’s definition of vulnerability were utilized. The assessment was based on various secondary data, like socio-economic and biophysical indicators, collected from several authenticated sources; and the respective weightage of these indicators was assigned by using ‘Principal Component Analysis’. Vulnerability was calculated as the net effect of exposure and sensitivity on the adaptive capacity. Anjaw district of eastern Arunachal Pradesh was found to be the most vulnerable district, while Tawang district of western Arunachal Pradesh happened to be the least vulnerable. This net effect was found negative in 7 out of 12 districts viz. Anjaw, Upper Siang, West Siang, Lower Dibang Valley, East Siang, East Kameng and Kurung Kurmey. This net negative effect could be construed as an alarming situation.

Yak (Bos grunniens) is a unique bovine species and considered as lifeline of highlanders. The male subfertility in yak is a matter of concern that causes huge economic loses. The spermatogenesis and male reproduction machinery are critically governed by Y-linked genes which tend to acquire necessary information in the course of evolution. The Y-linked fertility genes are present in multiple copies with testis-limited expression. To understand this novel complexity, 12 male-specific region of Y chromosome (MSY) genes have been studied in the yak. Targeted genes are amplified in male and female genomic DNA and confirmed the male derived specificity. Moreover, testis and sperm-specific expressions of MSY genes are distinct among different tissues. The quantitative polymerase chain reaction results validate the expression pattern of these genes in various tissues with predominant expression intestis and sperm. The sequencing of resultant yak MSY genes gives significant result and shows similarity with cattle (Bos indicus), but few nucleotide mismatches define the proposition of infertile male in the F1 hybrid of cattle and yak. The identified MSY genes can be used to establish male-specific characteristics and to differentiate male and female yak genotypically. Further, these genes may act as valuable resources to understand the capacity of spermatogenesis, embryogenesis, cellular growth, azoospermia and malesubfertility in the yak.

Yak, an economically important bovine species considered as lifeline of the Himalaya. Indeed, this gigantic bovine is neglected because of the scientific intervention for its conservation as well as research documentation for a long time.

Yak, an economically important bovine species considered as lifeline of the Himalaya. Indeed, this gigantic bovine is neglected because of the scientific intervention for its conservation as well as research documentation for a long time. Amelogenin is an essential protein for tooth enamel which eutherian mammals contain two copies in both X and Y chromosome each. In bovine, the deletion of a fragment of the nucleotide sequence in Y chromosome copy of exon 6 made Amelogenin an excellent sex-specific marker. Thus, an attempt was made to use the gene as an advanced molecular marker of sexing of the yak to improve breeding strategies and reproduction. The present study confirmed that the polymerase chain reaction amplification of the Amelogenin gene with a unique primer is useful in sex identification of the yak. The test is further refined with qPCR validation by quantifying the DNA copy number of the Amelogenin gene in male and female. We observed a high level of sequence polymorphisms of AMELX and AMELY in yak considered as novel identification. These tests can be further extended into several other specialized fields including forensics, meat production and processing, and quality control.

The molecular diversity of rumen methanogens was investigated using 16S rDNA gene library prepared from the rumen contents of Nili-Ravi buffaloes. Microbial genomic DNA was isolated from four adult male fistulated buffaloes and PCR conditions were set up using specific primers. Amplified product was cloned into a suitable vector, and the inserts of positive clones were sequenced. A total of 142 clones were examined, and the analysis revealed 46 species level (0.01 distance) operational taxonomic units (OTUs). Twenty six OTUs comprising 89 clones (63% of the total clones) were taxonomically assigned to Methanobacterium genus and the majority of them had highest percent identity with Methanobacterium flexile among cultured methanogens. Five OTUs comprising 27 clones (19% of total clones) were taxonomically assigned to Methanomicrobium genus and these clones showed highest sequence identity with Methanomicrobium mobile. Only two OTUs comprising 6 clones (4% of total clones) were assigned to Methanobrevibacter genus. A total of 17 clones belonging to 10 species level OTUs showed highest percent identity (ranging from 85 to 95%) with Methanomassilicoccus luminyensis and were taxonomically classified as Methanomassiliicocaceae. Out of the 142 rDNA clones, 112 clones, which constitute 79% of the total clones representing 42 OTUs, had less than 98.5% sequence identity with any of the cultured strains of methanogens and represent novel species of methanogens. This study has revealed the largest assortment of hydrogenotrophic methanogen phylotypes ever identified from the rumen of Nili-Ravi buffaloes. The study indicates that Methanobacterium is the most dominant methanogen in the rumen of Nili-Ravi buffalo. This is also the first report on the presence of methanogens phylogenetically close to M. luminyensis, an H2 dependent methylotrophic methanogen, in the rumen of buffaloes at such a high level of abundance.

In the present study, we report the distribution of true to type and atypical Nili-Ravi buffalo, a vulnerable dairy type riverine breed of North India and its underlying genetic structure. Out of total investigated buffaloes 73.5% had bilateral wall eyes while 5.4% had unilateral wall eyes and 21.1% had no wall eyes. 41.15% of Nili-Ravi buffaloes maintained in the breeding farm were having typical true to the type characteristics (both eyes walled, white markings in forehead, muzzle/chin, all the four legs and tail) while only 28.5% of Nili-Ravi buffaloes were true to the type under field conditions. Genotypic data were generated in four groups of Nili-Ravi buffalo (FMTNR--Typical Nili-Ravi from farm; FMANR--Atypical Nili-Ravi from farm; FDTNR--Typical Nili-Ravi from field; FDANR--Atypical Nili-Ravi from field) at 16 microsatellite loci. Comparative genetic analysis of various groups of Nili-Ravi buffaloes with Murrah revealed significant between group differences with an estimated global F(ST) of 0.063. Pair-wise F(ST) values ranged from 0.003 (between FDTNR and FDANR) to 0.112 (between FMTNR and FDTNR). Phylogenetic analysis and multi-dimensional scaling revealed clustering of FDTNR and FDANR together while FMTNR and FMANR clustered separately with Murrah in between farm and field Nili-Ravi buffaloes. Based on the results, the paper also proposes three pronged strategy for conservation and sustainable genetic improvement of Nili-Ravi buffalo in India.

Yak is an iconic symbol of Tibet and high altitudes of Northeast India. It is highly cherished for milk, meat, and skin. However, yaks suffer drastic change in milk production, weight loss, etc, when infested by parasites. Among them, infestation by leeches is a serious problem in the Himalayan belt of Northeast India. The parasite feeds on blood externally or from body orifices, like nasopharynx, oral, rectum, etc. But there has been limited data about the leech species infesting the yak in that region because of the difficulties in morphological identification due to plasticity of the body, changes in shape, and surface structure and thus, warrants for the molecular characterization of leech. In anticipation, this study would be influential in proper identification of leech species infesting yak track and also helpful in inventorying of leech species in Northeast India. Here, we investigated, through combined approach of molecular markers and morphological parameters for the identification of leech species infesting yak. The DNA sequences of COI barcode fragment, 18S and 28S rDNA, were analyzed for species identification. The generated sequences were subjected to similarity match in global database and analyzed further through Neighbour-Joining, K2P distance based as well as ML approach. Among the three markers, only COI was successful in delineating species whereas the 18S and 28S failed to delineate the species. Our study confirmed the presence of the species from genus Hirudinaria, Haemadipsa, Whitmania, and one species Myxobdella annandalae, which has not been previously reported from this region.

The cDNA encoding T cell receptor-zeta (TCR-ζ; CD247) molecule of water buffalo (Bubalus bubalis) was isolated, cloned and sequenced in the present study. The CD247 cDNA comprised 1078 nucleotides including a 30 nucleotide 5′-untranslated region (UTR), 495 nucleotide single open reading frame (ORF) and 553 nucleotide 3′-UTR. Deduced amino acid of buffalo CD247 sequence was two residues shorter than the corresponding cattle and sheep sequences. However, ruminant-specific insertions and substitutions in intra-cytoplasmic (IC) domain were present in buffalo. Immunoreceptor tyrosine-based activation motifs (ITAMs), the important motifs for TCR signalling, were totally conserved among ruminants including buffalo. The 3′-UTR region of the buffalo CD247 was highly homologous to the corresponding region in the cattle sequence and showed lack of polymorphism after polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) analysis using HaeIII and MseI restriction enzymes in buffalo population. ...

In the present study, water buffalo MHC (Bubu)-DRB cDNA was cloned and characterized. The 1022 base long-amplified cDNA product encompassed a single open reading frame of 801 bases that coded for 266 amino acids. The Bubu-DRB sequence showed maximum homology with the BoLA-DRB3*0101 allele of cattle. A total of seven amino acid residues were found to be unique for the Bubu-DRB sequence. The majority of amino acid substitutions was observed in the β1 domain. Residues associated with important functions were mostly conserved. Water buffalo DRB was phylogenetically closer to goat DRB*A.

The PCR- restriction fragment length polymorphism (RFLP) in and around TM4 of SLC11A1 gene and its association with the incidences of brucellosis in Hariana breed (Bos indicus) and Holstein Friesian crossbred (Bos indicusBos taurus) cattle was examined. A fragment of 954 bp encoding the TM4 was amplified, and RFLP was identified by digestion of the amplicon independently with AluI and TaqI. The amplicon (GenBank Acc. No. AY338470 and AY338471) comprised of a part of exon V (), and entire intron 5 (423 bp), exon VI (71 bp) and intron 6 (339 bp). Digestion with AluI revealed the presence of two alleles viz, A (281, 255, 79 and 51 bp) and B (541, 255, 79 and 51 bp). The frequency of A allele was estimated as 0.80 and 0.73 in Hariana and crossbred cattle, respectively. Due to presence of a polymorphic TaqI site at intron 5, two alleles: T (552 and 402 bp) and Q (231, 321 and 402 bp) were identified. The frequency of T allele was estimated as 0.96 and 0.97, respectively. For association study, on the basis of serological tests and history of abortion, the animals were grouped into "affected" and "non-affected". However, no association could be established with the observed RFLPs.

Sharma, D., Niranjan, S.K., Kumar, S., Deb, S.M., Naskar, S., Sharma, A. and Mitra, A. 2010. Molecular characterization of bubaline integrin β2 (ITGB2) cDNA. J. Appl. Anim. Res., 37: 217–220. A 2310-bp long buffalo (Bubalus bubalis) cDNA sequence of Integrin b2 (ITGB2) gene was characterized. At nucleotide sequence level, buffalo ITGB2 exhibited 96–98% homologies with other ruminants like cattle, bison, sheep, goat and deer. In coding region, total thirty two nucleotide differences have been observed between buffalo and cattle sequences including 15 non-homologous substitutions. Importantly, a conserved site (22) in PSI domain of the ruminants (Thr) was substituted in buffalo (Ala) indicating normal function of the integrin unless the second mutation in CD lie molecule coexists. Buffalo ITGB2 has seven differences in I-like domain region as compared to cattle. The transmembrane and cytoplasmic regions were highly conserved among buffalo and other ruminants. High homology and close phylogenetic as...

Panigrahi, M., Kumar, S., Deb, S.M., Mitra, A., Sharma, A. and Bujarbaruah, K.M. 2009. Lack of polymorphism in partial insulin like growth factor 1 (IGF1) and insulin like growth factor binding protein 3 (IGFBP3) genes of mithun. J. Appl. Anim. Res., 36: 41–44. A 396 bp fragment encompassing exon 5 of Insulin like growth factor 1 (IGF1) gene and 652 bp fragment encompassing exon 2–3 of Insulin like growth factor binding protein 3 (IGFBP3) gene were digested by HindIII and NlaIII restriction enzymes, respectively, in 90 mithun (Bos frontalis). The PCR-RFLP analysis showed the absence of polymorphism in these fragments in the animals screened. HindIII restriction enzyme produced two fragments of 269 and 127 bp in exon-5 of IGF 1. Similarly NlaIII restriction enzyme produced two fragments of 551 and 101 bp in exon 2–3 ofIGFBP3 gene. Two single nucleotide differences in IGF1 while six such differences in IGFBP3 gene existed in mithun sequences when compared to cattle. The sequence of the amplicons, w...

A fragment of 570 bp corresponding to exon 5 and 6 of integrin beta 2 (ITGB2) gene was amplified for screening D128G mutation in one hundred and fifty two buffaloes (Bubalus bubalis) which causes bovine leukocyte adhesion deficiency syndrome (BLAD) in cattle, as well as to ascertain polymorphism. TaqI PCR-RFLP revealed no such mutation thus indicating the absence of bubaline leukocyte adhesion deficiency (BuLAD) allele in animals under study. However, the polymorphism studies using MspI restriction enzyme revealed two genotypic patterns viz. AA pattern (bands of 293, 141, 105, and 31 bp) and BB pattern (bands of 293, 105, 77, 64, and 31 bp). The sequences of A and B alleles were submitted to the GenBank (EU853307 and AY821799).

Kumar, S., Gupta, S., Manohar R.P.V., Deb, S.M., Mitra, A., Sharma, A. and Bujarbaruah, KM. 2009. Nucleotide variability of partial somatotropin hormone (STH) gene in mithun (Bos frontalis). J. Appl. Anim. Res., 35: 165–168. A fragment of 544 bp corresponding to exon 1 and 2 of somatotropin hormone (STH) gene was amplified in 90 mithun (Bos frontalis). The nucleotide sequence similarity of mithun with zebu cattle, yak, exotic cattle and buffalo was found to be 99.6, 99.4, 98.9 and 93.5%, whereas, the corresponding figures for deduced amino acid sequence were 100, 100, 99.4 and 98.8%, respectively. The PstI and PvuII PCR-RFLP analysis produced bands of 479, 65 and 448, 96 bp, respectively, in all the animals indicating no mutant allele and hence monomorphism in the mithun population studied. The sequence of the amplicon was submitted to GenBank (Accn no. EU166347).

John, B., Kumar, S., Deb, S.M., Mitra, A., Niranjan, S.K., Naskar, S. and Sharma, A. 2006. Genetic variability in Integrin Beta-1 (ITGB1) gene of buffaloes. J. Appl. Anim. Res., 30: 149–152. A fragment of 151 bp corresponding to exon-5 of ITGB1 gene was amplified in 45 buffaloes (Bubalus bubalis) of murrah breed. The RFLP analysis showed the absence of polymorphism in this fragment with respect to Trull restriction enzyme and showed only one restriction site, which produced two fragments of 21 and 130 bp. The putative metal ion binding D-X-S-X-S sequence motif in the I-like domain of b1 subunit was highly conserved in these buffaloes. Three and nine nucleotide sequence variations were found in buffalo sequences when compared to that of cattle and human, respectively. The sequence of the amplicon which was the first report on buffalo ITGB1, was submitted to GenBank.

Anbarasan, K., Sharma, A.K., Singh, R.K., Deb, S.M. and Sharma, D. 2001. Genetic differentiation among goats using randomly amplified polymorphic DNA (RAPD) markers. J. Appl. Anim. Res., 20: 83–88. To develop population specific markers using randomly amplified polymorphic DNA—ploymerase chain reaction (RAPD—PCR) in meat (Black Bengal and Non-descript Rohilkhand Local), milk (Barbari) and pashmina (Chegu) producing goats, a total of 35 random sequence decamer primers were tested. Nine primers generated polymorphism in one or more goat populations. Out of 117 bands scored, 46 (39%) were observed to be polymorphic. One of the primers (OPM-18) was found effective in differentiating the Pashmina (Chegu) from meat or milk type of goats. While the primer OPI-05 generated one band which might be specific for meat type goats i.e. Black Bengal and Non-descript (Local). However, population specificity and sequence homology for these bands need further investigation by hybridization, cloning and sequencing ...

Kumar, S., Manohar, R.P.V., Deb, S.M., Mitra, A., John, B., Sharma, A. and Bujarbaruah, K.M. 2008. Genetic variability of somatotropin releasing hormone (SRH) gene in Mithun (Bos frontalis). J. Appl. Anim. Res., 33: 205–207. A fragment of 452 bp corresponding to exon 2 and 3 of somatotropin releasing hormone (SRH) gene on amplification in 90 mithun (Bos frontalis) revealed two genotypie patterns viz. AA genotypie pattern (bands of 292 and 160 bp) and AB genotypie pattern (452, 292, and 160 bp) with respect to Saul restriction enzyme. The sequence of the A allele, which was the first report on mithun SRH gene, was submitted to GenBank (Accession No. EF014289).

Niranjan, S.K., Deb, S.M., Mitra, A., Sharma, D., Sharma, A and Sharma A. 2005. Partial genomic sequence of buffalo (Bubalus bubalis) ITGB2 gene. J. Appl. Anim. Res., 28: 41–43. A fragment of 367 bp corresponding to parts of fifth and sixth exons flanking intron five of ITGB2 was amplified in river buffaloes (Bubalus bubalis). The Taq I-RFLP analysis showed the absence of BLAD (D128G) allele in the buffalo population studied. The sequence data of the amplicon, first report on buffalo ITGB2, was submitted to Genbank. Single nucleotide changes at seven places in fifth exon of buffalo ITGB2 led to substitution of Val140 and Asp159 and creation of one Msp I site.

A study was carried out (375 days) in the outer Himalayas (29 °25′N and 79 °35′E) where the agroclimate for the major part of the year is cold and humid. Nine male pashmina (Cheghu) kids, (aged 5–6 months) with mean body weight 11.3 ± 0.5 kg, were randomly assigned to three groups of three animals each in a completely randomized design. Animals were fed a concentrate mixture along with ad lib. oats hay (Group I) or freshly lopped green oak (Quercus semecarpifolia) leaves (Groups II and III), while Groups I and II were provided CP and TDN requirements (as per NRC recommendations), animals in Group III were given 16% higher CP. A metabolism trial was conducted 270 days post experimental feeding. DM intake, average daily gain (ADG), pashmina fibre yield, urinary N excretion, N balance, N retention as % of N absorbed, rumen liquor pH and TCA soluble N did not differ significantly among groups. Digestibility of DM, CP, EE, NDF and ADF was significantly depressed in Group II as compared with Group I. Between Groups I and III, digestibility of CP and EE was comparable but DM, NDF and ADF digestibilities were higher (P 0.05) in all groups. Between Groups I and III, mean values of VFA and various N fractions in strained rumen liquor were identical except TCA ppt. N, which was increased (P < 0.01) in Group I. No lesions of any significance were found in the vital organs of the animals in the various groups. The depressing effect of oak leaves on digestibility of nutrients could be ameliorated by providing higher CP content in the diet of animals. It may, therefore, be concluded that 25–30% CP in the diet of pashmina goats can safely be furnished by oak leaves without affecting their health and production.

The genetic diversity of MHC class II DQ genes was investigated in riverine buffalo (Bubalus bubalis) by PCR-RFLP and sequencing. Highly variable regions (exons 2-3) of DQ genes were amplified from 152 buffaloes and genotyped by PCR-RFLP. Alleles identified by differential restriction patterns were sequenced for the characterization. PCR-RFLP was a rapid method to discriminate between DQA1 and duplicated DQA2 genes in buffalo, however, the method appeared to be inadequate for determining the more complicated DQB genotypes. A total of 7 and 10 alleles were identified for DQA and DQB loci, respectively. Nucleotide as well as amino acid variations among DQ alleles particularly at peptide binding regions were high. Such variations were as expected higher in DQB than DQA alleles. The phylogenetic analysis for both genes revealed the grouping of alleles into two major sub-groups with higher genetic divergence. High divergence among DQ allelic families and the isolation of two diverse DQA and DQB sequences from individual samples indicated duplication of DQ loci was similar in buffalo to other ruminants.

In the present study, we explored structural and functional variations and possible duplication of the major histocompatibility complex (MHC)-DQA gene in water buffalo (Bubalus bubalis). Two cDNA sequences, amplified from one individual water buffalo, were designated as Bubu-DQA1 (DQA*0101) and -DQA2 (DQA*2001). The percentage of nucleotide and amino acid similarity between Bubu-DQA1 and -DQA2 revealed that these sequences display more similarity to alleles of respective DQA1 and DQA2 genes from other ruminant species than to each other. The phylogenetic analysis also revealed a considerably larger genetic distance between these two genes than between homologous genes from other species. The larger genetic distance between DQA*0101 and DQA*2001, and the presence of different bovine DQA putative locus specific amino acid motifs, suggests these sequences are non-allelic. This finding is consistent with DQA gene duplication in other ruminants.

Yak is one of the most important economically useful animals for highlanders. The decline in the yak population demands effective measures for conservation and multiplication of elite germplasm. In vitro production of embryos and their cryopreservation and transfer to suitable recipients for production of elite calves may contribute to fulfill the objectives. The work was conducted at the National Research Center on Yak over a period of 3 years. The ovaries of slaughtered animals were used for collecting oocytes through aspiration of follicles followed by slicing of ovaries in the conventional method. Trials were conducted using 7 cyclic parous yaks for ultrasound-guided ovum pickup (OPU) at Nyukmadung farm (2700 m above mean sea level). The technique followed was similar to that in buffaloes with slight modification. Categories of oocytes classified A (2–3 layers of cumulus) and B (at least one layer of cumulus) obtained through the processes were subjected to in vitro maturation using standardized maturation medium (TCM-199 + 10% follicular fluid + sodium pyruvate + l-glutamine + 10% heat inactivated oestrus cow serum + pFSH + 17β oestradiol). The frozen-thawed yak sperm were capacitated using the swing-up method before their incubation with matured oocytes using BO medium. Oocytes matured for 24 h were washed 5 to 6 times with BO medium and then co-incubated with in vitro capacitated spermatozoa (0.1 to 0.25 million) for fertilization (8–10 oocytes per group) in 100-µL droplets of BO medium under mineral oil in 35-mm Petri dishes and placed in a CO2 incubator (5% CO2, 90% RH) at 38.5°C for 16 to 18 h. The presumed zygotes were washed several times in mCR2aa (modified Charles Rosenkrans) washing medium and then cultured in culture medium for 7 days on original beds of granulosa cells. The rates of maturation and fertilization of oocytes collected by conventional and OPU technique were comparable (Table 1). This may be attributed to greater numbers of good quality oocytes recovered in the conventional method. Embryos developed up to the stage of compact morula and blastocysts (24.66% through conventional and 22.73% through OPU) were cryopreserved using the vitrification method for further study. Thirteen embryos were transferred non-surgically to one each of 13 yak recipients; 5 became pregnant and only 1 recipient transferred with a cryopreserved-thawed embryo, developed through OPU, delivered one male calf, leading to the first successful production of an IVF yak calf in the world. The present findings are suggestive of using the OPU technique for in vitro embryo production, though resulting in lower numbers of transferable embryos (Table 1), because availability of ovaries for conventional IVF is a major constraint in yak. Table 1.Comparative in vitro yak embryo production rate with recovery of oocytes by conventional or ovum pickup (OPU) method

Natural resistance associated macrophage protein 1 ( NRAMP1 ), an integral transmembrane protein, is reported to influence the intraphagosomal microbial replication and thereby confer resistance to several intracellular pathogens in mice. In bovine, a significant association of (GT) 13 allelic variant of polymorphic microsatellite at 3′ untranslated region (UTR) of NRAMP1 gene with natural resistance to brucellosis has been established. The present study was aimed to detect polymorphism at 3′UTR of NRAMP1 gene in Hariana breed of Bos indicus cattle and Holstein Friesian crossbred ( B. indicus × Bos taurus ) cattle, and to determine the association of this polymorphism with resistance/susceptibility to brucellosis. The (GT) n polymorphism at 3′UTR in terms of variation in fragment length was determined using denaturing polyacrylamide gel analysis of radioisotope incorporated amplicon of 174 bp. Screening of a total of 100 samples (comprising 50 random samples of each breed) revealed that animals were of same genotype, i.e., homozygous (GT) 13 /(GT) 13 . Sequencing of amplicons from representative animals confirmed the presence of (GT) 13 repeat. For association study, the animals that were positive in all three serological tests (viz., RBPT, STAT and ELISA) and had history of abortion were grouped as “affected”; whereas the animals that were negative in all these tests and completed third lactation without any history of abortion were grouped as “non-affected”. Since, all animals belonging to either group were homozygous (GT) 13 , association could not be established. However, the present study demonstrated that the presence of (GT) 13 allele even in homozygous condition could not provide enough resistance to brucellosis in a naturally infected herd.