Caudal anaesthesia of the infraorbital nerve for diagnosis of idiopathic headshaking and caudal compression of the infraorbital nerve for its treatment,in 58 horses

Reasons for performing study: Idiopathic headshaking is often a facial pain syndrome, but a diagnostic protocol has not been described. In a previous study, caudal compression of the infraorbital nerve for treatment offered a fair success rate, but low case numbers and short follow‐up time were limitations. Objectives: To describe a diagnostic protocol for headshaking, examining the role of bilateral local analgesia of the posterior ethmoidal nerve (PET block). To report longer‐term follow‐up after surgery of the original cases and further cases and to determine whether changes to the technique influence success rates and complications. Methods: Records of horses that had undergone PET block and caudal compression surgery at 3 hospitals were reviewed. Modifications to the surgical technique included placing additional coils into the infraorbital canal and/or performing concurrent laser cautery of the nerve. Follow‐up information was obtained by telephone contact with owners. Results: The PET block was performed in 27 horses, with a positive result in 23 of 27 (85%). Surgery was performed in 58 horses. A successful outcome was initially achieved in 35 of 57 (63%) horses, but recurrence occurred between 9 and 30 months later in 9 (26%). Surgery was repeated in 10 of 31 (32%) horses. Final success rate, considering only response to the last performed surgery, was 28 of 57 (49%) horses with median follow‐up time of 18 months (range 2–66 months). Nose‐rubbing was reported post operatively in 30 of 48 (63%) horses. This resolved in all but 4 horses, which were subjected to euthanasia. Response to PET block or change in surgical technique did not appear to influence outcome or complications. Conclusions and potential relevance: The diagnostic protocol described is recommended for the investigation of headshakers. Caudal compression offers the best prognosis for a successful outcome compared with other treatments, for horses in which the only alternative is euthanasia. Surgical treatment of the disorder requires refinement, and the pathogenesis of the disorder requires investigation.     

m‐carboxycinnamic acid bishydroxamide improves developmental competence,reduces apoptosis and alters epigenetic status and gene expression pattern in cloned buffalo (Bubalus bubalis) embryos

Incomplete or aberrant reprogramming of nuclear genome is one of the major problems in somatic cell nuclear transfer. In this study, we studied the effect of histone deacetylase inhibitor m‐carboxycinnamic acid bishydroxamide (CBHA) on in vitro development of buffalo embryos produced by Hand‐made cloning. Cloned embryos were treated with CBHA (0, 5, 10, 20 or 50 μM) for 10 hr from the start of reconstruction till activation. At 10 μM, but not at other concentrations examined, CBHA increased (p < .05) the blastocyst rate (63.77 ± 3.97% vs 48.63 ± 3.55%) and reduced (p < .05) the apoptotic index of the cloned blastocysts (8.91 ± 1.94 vs 4.36 ± 1.08) compared to untreated controls, to levels similar to those in IVF blastocysts (4.78 ± 0.74). CBHA treatment, at all the concentrations examined, increased (p < .05) the global level of H3K9ac in cloned blastocysts than in untreated controls to that observed in IVF blastocysts. Treatment with CBHA (10 μM) decreased (p < .05) the global level of H3K27me3 in cloned blastocysts than in untreated controls but it was still higher (p < .05) than in IVF blastocysts. CBHA (10 μM) treatment increased (p < .05) the relative expression level of pluripotency‐related genes OCT‐4 and NANOG, and anti‐apoptotic gene BCL‐XL, and decreased (p < .05) that of pro‐apoptotic gene BAX than in untreated controls but did not affect the relative expression level of apoptosis‐related genes p53 and CASPASE3 and epigenetics‐related genes DNMT1, DNMT3a and HDAC1. These results suggest that treatment of cloned embryos with 10 μM CBHA improves the blastocyst rate, reduces the level of apoptosis and alters the epigenetic status and gene expression pattern.     

Relationship between endometritis and oxidative stress in the follicular fluid and luteal function in the buffalo

In this study, alteration in the follicular fluid composition and luteal function was investigated in the buffalo with endometritis. Genitalia were classified into cytological and purulent endometritis on the basis of polymorphonuclear cell cut off while non‐endometritis served as control (n = 10/group). In the follicular phase, the number of surface follicles was counted, diameter of the largest follicle was measured and the follicular fluid was assayed for total protein, cholesterol, malondialdehyde (MDA), total antioxidant capacity (TAC), oestradiol (E₂) and progesterone (P₄). The P₄ content of corpus luteum during mid‐luteal phase was estimated by radioimmunoassay. Ovaries from the follicular phase of oestrous cycle showed no significant difference in the total number of surface follicles, size of the largest follicle and volume of follicular fluid in the buffaloes with and without endometritis (p > .05). However, the antral fluid of the largest follicle from the genitalia of buffalo with cytological and purulent endometritis showed a significant decrease in the concentration of total protein, cholesterol, TAC and E₂ and a significant increase in the concentration of MDA and P₄ (p < .05). The results indicated that there is an association between endometritis and decreased ovarian function.     

Detection and localization of regucalcin in spermatozoa of water buffalo (Bubalus bubalis): A calcium‐regulating multifunctional protein

Regucalcin (RGN) is a calcium‐regulating, anti‐apoptotic, antioxidative and antiproliferative multifunctional protein predominantly seen in liver and kidney. All these functions are very crucial during spermatogenesis and sperm maturation process until fertilization of the ovum. Although many studies have reported the wide distribution of regucalcin in the male reproductive tract of the rat, human and bovine, its presence in spermatozoa is yet to be demonstrated wherein calcium has a pivotal role in the transport, capacitation, acrosomal reaction and further fusion with ova. Here, we detected the expression of regucalcin mRNA and protein in buffalo spermatozoa using real‐time PCR and Western blot, respectively. The study detected two new regucalcin isoforms of 44 kDa and 48 kDa size along with the reported 34‐kDa, 28‐kDa and 24‐kDa isoforms, wherein the 34‐kDa isoform was found to be membrane associated in spermatozoa. Further, immunocytochemistry study localized the regucalcin protein in the acrosomal region of the caudal and ejaculated buffalo spermatozoa while it was detected in both cytoplasm and acrosomal region of testicular spermatozoa. This discovery of RGN in spermatozoa and localization in the acrosomal region will help to focus researchers to see its role in calcium‐related functions like capacitation, acrosomal reaction and membrane fusion. Overall, regucalcin may be a new fertility marker in buffalo and can be utilized for infertility treatments.     

Antisperm antibodies in repeat‐breeding cows: Frequency,detection and validation of threshold levels employing sperm immobilization,sperm agglutination and immunoperoxidase assay

Antisperm antibodies have been found in repeat‐breeding(RB) cows, and those causing agglutination and/or immobilization of sperm are considered to be closely related to unexplained infertility. However, a standard protocol for identifying antisperm antibodies (ASA) in cattle is not validated. Therefore, an investigation was undertaken to evaluate sperm immobilization (SIT), sperm agglutination (SAT) and immunoperoxidase (IPT)assays for detection of ASA in serum and their respective threshold levels for confirmation. Animals (heifers, normally breeding, repeat‐breeding and pregnant animals) that were free from IBR, brucellosis and uterine infections (screened by clinical examination) were included in the study. Sperm agglutinating, sperm immobilizing and antisperm antibodies evaluated by respective assay were significantly higher (p < .05) in RB cows compared to other groups. The SIT assay was able to identify 61% of RB caused by ASA, more than those employing SAT and IPT. Furthermore, a dilution rate of 1:5 and 1:80 (confirms 59.0 and 57.0% RB+ve)were sufficient to diagnose ASA by SAT and IPT, respectively. Results indicate the presence of __12.6% clumped spermatozoa and __ 2.6%(cut‐off value) peroxidase‐positive spermatozoa at 1:5 and 1:80 dilutions diagnosed with SAT and IPT, respectively, may be considered as repeaters arising out of ASA. Furthermore, study also showed the presence of lower incidence of ASA positivity in other groups of animals (heifer<normal breeder<pregnant animals) compared to repeaters. Study results show that although IPT is more specific and accurate but SAT and SIT are comparatively simple and cost‐effective assays suitable for detecting ASA under field conditions and thus can be recommended for screening of repeaters.     

Conventional and contemporary approaches for drought tolerance rice breeding: Progress and prospects

Drought is becoming a major threat to rice farming across the globe owing to the depletion of water tables in rice-growing belts. Drought affects rice plants at multiple stages, causing damage at morphological and physio-biochemical levels, leading to severe losses that exceed losses from all other stresses. The amalgamation of conventional breeding methods with modern molecular biology tools and biometrical methods could help accelerate the genetic gain for drought tolerance in rice. Many drought-tolerance traits with genetic determinants have been identified and exploited for tolerance rice variety breeding. The integration of genome-wide association study and genomic selection tools with speed breeding shortened the breeding cycle and aided in rapid improvement of genetic gain. In this review, we emphasized the progress made through classical breeding as well as the limitations and usefulness of current genomic methods in improving drought tolerance. We briefly addressed methods for identifying genetic determinants for drought tolerance and deploying them through genomics-assisted breeding programmes to develop high-yielding drought-tolerant rice cultivars.     

Draft Genome Sequence of Potato Dry Rot Pathogen <Emphasis Type="Italic">Fusarium sambucinum</Emphasis> Fckl. F-4

Fusarium sambucinum is one of the most important causal agents that not only cause the dry rot disease of potato tubers in fields and stores worldwide but also capable of producing secondary metabolites toxic for people and animals. Here we present the first draft genome sequence of the strain (F-4) estimated to be around appx. 42.0 Mb. The genome has 12,845 protein coding genes with more than 35,900 exons and gene density of 3.13 per 10Kb. F. sambucinum is evolutionary more close to the F. graminearum among the Fusarium species complex. The genome sequence represents a valuable resource for understanding the pathogenecity and virulence factors, and their evolution within the complex and highly plastic genus Fusarium.     

Detection,Localization and Tyrosine Phosphorylation Status of Ser/Thr Protein Phosphatase1γ in Freshly Ejaculated,In Vitro Capacitated and Cryopreserved Buffalo Spermatozoa

Several recent studies have indicated the important roles of Ser/Thr protein phosphatase1γ (PP1γ) in regulating the motility and capacitation of mammalian spermatozoa. Here, we report the presence and distribution of PP1γ protein in freshly ejaculated, in vitro capacitated and cryopreserved buffalo spermatozoa. The presence of PP1γ and its distribution were assessed by Western blotting and indirect immunofluorescence techniques, whereas the isoforms of PP1γ and their tyrosine phosphorylation status were identified by using 2D electrophoresis. The number of isoforms and the status of tyrosine phosphorylation of PP1γ were increased in capacitated spermatozoa when compared with freshly ejaculated spermatozoa. Differential pattern of expression and tyrosine phosphorylation of PP1γ were observed in cryopreserved spermatozoa, wherein some isoforms were degraded and some were tyrosine phosphorylated. In addition, immunofluorescence technique revealed that PP1γ was localized to principle, mid‐piece, post‐acrosomal and equatorial regions of buffalo spermatozoa. Differential distribution of tyrosine‐phosphorylated proteins were observed in fresh, capacitated and cryopreserved spermatozoa. The tyrosine phosphorylation of several proteins (20, 37, 38, 52, 60, 79 and 100 kDa) were increased when sperm cells were incubated with PP1γ inhibitor, okadaic acid. Together, our results suggest that buffalo spermatozoa express different isoforms of PP1γ protein. The protein expression and tyrosine phosphorylation of PP1γ were increased during capacitation. Furthermore, the differential pattern of expression and tyrosine phosphorylation of PP1γ were observed in cryopreserved spermatozoa. In addition, the inhibition of PP1γ protein increases protein tyrosine phosphorylation in capacitation.     

不同种植密度和施肥水平对大豆籽粒品质的影响

本试验自2002年开始至2004年止，从种植密度和施肥水平两方面对大豆脂肪、蛋白质含量的影响进行了分析。结果表明：种植密度对大豆籽粒的脂肪、蛋白质含量具有显著的影响，相关性分析结果显示，种植密度与大豆籽粒的脂肪含量存在着负相关性，与籽粒中的蛋白质含量之间为正相关。中国品种的脂肪含量受种植密度的影响大于美国品种，而中国品种的蛋白质含量受种植密度的影响小于美国品种。蛋脂总含量受种植密度的影响较小，并且不存在品种间的差异性。磷酸二铵的施用量与大豆籽粒脂肪含量和蛋白质含量之间没有显著的相关性，但是中美品种的品质对磷酸二铵施用量的反应不同。磷酸二铵的施用量与中国品种的脂肪含量、蛋白质含量和蛋脂总含量呈正相关，而与美国品种的脂肪、蛋白质含量和蛋脂总含量呈负相关，但相关程度均不显著。