Seven-week asynchronous twins in a buffalo heifer (Bubalus bubalis) associated with superfetation – A case report

A female buffalo (Bubalus bubalis) of the Bulgarian Murrah breed aged 1,090 days was observed to give birth to a second newborn (normally developed male) after she had calved (normal female) 49 days earlier. This phenomenon is highly associated with her melatonin treatment within a trial for induction of puberty, the last ear implants being placed approximately 50 days before the assumed date of first mating, to which point the level of progesterone had increased dramatically. Despite none of the matings of the dam was visually witnessed to prove ovulation over an existing gestation, we take the liberty to qualify this phenomenon as superfetation, ruling out the other possible phenomena, namely embryonic diapause as it is highly unlikely to occur in any livestock species, and differentiated development of twin foetuses as it is associated with foetal malformation, which was not observed in this case.     

Multiple domestication of swamp buffalo in China and South East Asia

The domestication of swamp buffalo (Bubalus bubalis carabanesis) has been discussed for years and still remained unclear. To obtain a better understanding of where, when and how the swamp buffaloes were domesticated, the complete mitochondrial DNA D-loop sequences of 1,788 individuals from China, Vietnam, Laos, Thailand, Burma, Bangladesh and India were investigated. Our results revealed swamp buffalo with abundant genetic diversity. The lineage SA of swamp buffalo may be first domesticated in Lower Yangtze and then migrated following two different routes: one migrated along the Upper Yangtze to Southwest China (L1); the other migrated to Southeast China to Southeast Asia through Guangxi province (L2). During the migration process several later domestication events may be inspired in the Upper Yangtze (SB3), Southwest China and North of Southeast Asia (SB1 and SB2). In this study, we proposed a non-independent multiple domestication pattern in swamp buffalo.     

人神经母细胞瘤细胞异种克隆胞质受体的选择

本研究检测人神经母细胞瘤细胞-水牛和人神经母细胞瘤细胞-猪重构胚胎的体外发育潜能,以期建立人神经细胞母细胞瘤细胞的异种治疗性克隆模型,为进一步研究人癌症发生过程中的表观遗传学修饰机制及分离得到正常的人类胚胎干细胞奠定基础。分别以水牛颗粒细胞和人神经母细胞瘤细胞为供体,水牛卵母细胞为受体构建克隆胚胎,然后检测这2种重构胚胎的体外发育效率;再以猪卵母细胞为胞质受体,同样以上述2种体细胞为供体细胞构建重构胚胎。结果显示,人神经母细胞瘤细胞-水牛重构胚胎囊胚发育率显著低于水牛同种克隆胚胎的（1.1%∶12.1%,P〈0.05）,而融合率（83.7%∶79.4%,P〉0.05）和卵裂率（55.8%∶58.2%,P〉0.05）无显著差异。但是,当以猪卵母细胞为受体胞质时,人-猪重构胚胎卵裂率显著低于水牛-猪重构胚胎（43.7%∶89.8%,P〈0.05）,但重构胚胎融合率和囊胚率差异不显著（85.8%∶82.5%,0∶1.4%,P〉0.05）。结果表明,人神经母细胞瘤细胞-水牛和人神经母细胞瘤细胞-猪异种重构胚胎可以在体外发育,尽管其囊胚发育率相对比较低。     

槟榔江水牛保种及选育利用策略

槟榔江水牛是近年在云南西部发现的我国第一个本土河流型水牛群体,具有较高的产奶性能,是发展我国水牛奶业宝贵的遗传资源。本文基于近几年对槟榔江水牛遗传资源调查和种质特性及遗传背景分析的结果,就其遗传资源的保护和选育利用方略进行了探讨。     

Single Nuclease Polymorphism Analysis of Leptin Gene in Buffalo

This study was aimed to detect the polymorphisms of Leptin gene in buffalo that provided a fundamental for further study on marker assisted breeding in buffaloes.The single nuclease polymorphisms (SNP) of Leptin gene were identified and genotyped by using DNA pooled sequencing and high-resolution melting (HRM) method in three buffalo breeds with 182 buffalo individuals,respectively.The results showed that seven SNPs of Leptin gene were identified in studied population that were located in the intron 1,intron 2 and exon 3 regions,respectively.All the SNPs loci were moderate polymorphism except for the SNP5 and SNP6.The χ²-test indicated that all the SNPs loci were in agreement with Hardy-Weinberg equilibrium in Nili-Ravi population (P>0.05).Our findings revealed that seven SNPs of Leptin gene in buffalo were identified and genotyped in population,which provide the data support for further analyzing the associations between these polymorphism and production traits in buffaloes.     

Cloning,Analysis and Construction of Eukaryotic Expression Vector of Buffalo MBD3 Gene

In this study,through cloning buffalo MBD3 gene and analyzing the biological information of MBD3 gene sequence,and constructing the expression vector of buffalo MBD3,to provide a basis for the function research of buffalo MBD3 gene on embryo development and iPCSs.The total RNA was extracted from buffalo fresh ovary,and MBD3 gene was amplified and sequenced,the sequence was systemically analysed with bioinformatics techniques.And the MBD3 CDS was cloned into the pEGFP-C1 vector.Then the recombinant plasmid pEGFP-C1-MBD3 was transferred into the HEK293T cells and buffalo fetal fibroblasts (BFF),the expression was analyzed by RT-PCR,Western blotting and fluorescence microscope.The results showed that 898 bp of MBD3 gene fragment including whole 774 bp CDS was cloned and sequenced,and encoded 257 amino acids.The multiple sequence alignment and analysis of phylogeny tree showed that MBD3 gene was highly conserved in the process of evolution.Especially the MBD domain,the MBD domain of buffalo MBD3 gene shared 100% of similar nucleotide sequence with Bos taurus,and shared 97% of similar nucleotide sequence with Homo sapiens,Sus scrofa and Pan troglodytes.The recombinant plasmids pEGFP-C1-MBD3 were transferred into HEK293T cells and BFF,fluorescence observation,RT-PCR and Western blotting were used to analyze the expression of buffalo MBD3.The results suggested that the expression vector of buffalo MBD3 gene was successfully constructed.The study laid the foundation for the function research of MBD3 on embryo development and inducing of iPCSs.     

Exogenous cholesterol prevents cryocapacitation-like changes,membrane fluidity,and enhances in vitro fertility in bubaline spermatozoa

A study was conducted to determine the optimum dosage of the exogenous cholesterol-loaded cyclodextrins (CLC) to get maximum cryoprotection for bubaline spermatozoa. In the present study, 120 × 10⁶ spermatozoa were incubated in 2, 3 and 4 mg of CLC as grouped as Gr II, III and IV, respectively, and sperm progressive motility, intracellular Ca²⁺, capacitation status by protein tyrosine phosphorylation (PTP) assay and zona binding per cent (ZBP) and cleavage rate (CR) of the cryopreserved buffalo spermatozoa by in vitro fertility assay were assessed in comparison with an untreated control group (Gr I). Results revealed that there was a significant (p < .05) linear decrease in percentage of sperm population with higher intracellular Ca²⁺ and percentage of sperm population with medium or high capacitated by PTP in CLC treated from 2 to 3 mg and then increased to 4 mg/120 × 10⁶ spermatozoa whereas sperm progressive motility, percentage of sperm population with low capacitated, ZBP and CR were increased significantly (p < .05) in sperm population treated from 2 to 3 mg CLC and then decreased to 4 mg/120 × 10⁶ spermatozoa. The study has clearly indicated that CLC at 3 mg/120 × 10⁶ spermatozoa has maximum beneficial effects in protection of sperm progressive motility, membrane fluidity (low intracellular Ca²⁺); prevention of cryocapacitation (low capacitation pattern in immunolocalization) and enhancement of in vitro ZBP and CR. Post-thaw motility of the CLC-treated sperm has shown positively significant (p < .05) correlation with sperm population with low intracellular Ca²⁺, low capacitated sperm population, ZBP and CR, whereas it was negatively (p < .05) correlated with sperm population with high intracellular Ca²⁺, medium or high capacitated sperm. The present study has revealed for the first time that incubation of spermatozoa with CLC of higher dose (>3 mg/120 × 10⁶ spermatozoa) had adverse effects on sperm cryopreservation, although incubation of sperm with 3 mg/120 million prior to processing had minimised the freezing–thawing-associated damages in bubaline species.     

Comparison of carcass and meat characteristics of Brahman grade cattle (Bos indicus) and crossbred water buffalo (Bubalus bubalis)

This study was conducted to determine and compare the live traits, carcass yield and the physical, chemical, processing and sensory properties of meat from 20 Brahman grade cattle (crossbred cattle) and crossbred water buffalo (Philippine carabao × Murrah, Bulgarian or Indian) with an average age and weight of 29 months and 434 kg, respectively. The animals were fed a similar diet for 180 days before slaughter. A survey was performed to establish consumer meat preference between the two species. Live weights before slaughter of the two species were found to be different (P < 0.05) with the crossbred water buffalo showing higher bodyweight. However, the crossbred water buffalo had a lower (P < 0.05) dressing yield based on hot carcass weight and chilled carcass weight. The lower dressing yield of crossbred water buffalo can be attributed to its higher (P < 0.05) weights of edible and non‐edible slaughter by‐products. The chilled carcass yield of the forequarters and hindquarters from crossbred cattle and crossbred water buffalo were comparable when expressed as percent of live weight and chilled carcass weight. The estimated lean yield was higher in crossbred cattle than crossbred water buffalo. Crude protein, ash, fat, cholesterol, myofibrillar, sarcoplasmic and insoluble protein contents of the beef and carabao meat were all similar. Water holding capacity, pH, muscle fiber diameter, tenderness, firmness and marbling score in carabeef were all comparable to the beef. Redness was found to be higher (P < 0.05) in carabeef than the beef as measured with a color meter. The consumer meat preference survey showed that 55.9% selected the beef while 44.1% preferred carabeef. The color and amount of fat on the outside of the beef sample were primary in buyer selection.     

Follicular dynamics and colour Doppler vascularity evaluations of follicles and corpus luteum in relation to plasma progesterone during the oestrous cycle of Surti buffaloes

With an objective to evaluate the follicular dynamics and vascularity changes in follicles and corpus luteum, the ovaries of cyclic Surti buffaloes (n = 9) were examined daily sequentially by transrectal B‐mode and colour flow mode (CFM) ultrasonography starting from the day of oestrus till the onset of next oestrus. Higher proportion of buffaloes evidenced one‐wave cycle (66.66%) compared to two‐wave cycle (33.34%) with none showing a three‐wave cycle. The dominant follicle of the first follicular wave was the ovulatory follicle and persisted for 19.70 ± 0.50 days compared to its persistence for 16.5 ± 1.45 days in a two‐wave cycle. The maximum diameter of the ovulatory follicle in a one‐wave and two‐wave cycle did not differ yet their linear growth rates were significantly lower (p < 0.01) in a one‐wave cycle. Colour flow mode examination of follicles revealed that the percentage of follicles with detectable blood flow in the subsequently determined largest follicle (dominant follicle) was not different from that in the second largest follicle before follicle deviation. The blood flow in the dominant follicle increased significantly on the day of oestrus. The mean diameter and blood flow to the corpus luteum (CL) increased linearly and significantly from Day 5 of oestrus till Day 13 after which both parameters started declining. At or around Day 16, there was precipitous fall in the blood supply to the CL and CL diameter that continued declining thereafter to reach the lowest around Day 20 of the oestrous cycle. Rise in plasma progesterone concentrations was synchronous to CL diameter and vascularity and showed significant and positive correlations. It was concluded that Surti buffaloes evidence a preponderance of one‐wave follicular growth pattern with a significant increase in the vascularity of ovulatory follicle on the day of oestrus and corpus luteum on Day 13 of the oestrous cycle.