Cardiac arrhythmias are probably more common in horses than in any other domestic animal species where poor performance and exercise intolerance is the most frequent clinical complaint. Atrial fibrillation is a type o...Cardiac arrhythmias are probably more common in horses than in any other domestic animal species where poor performance and exercise intolerance is the most frequent clinical complaint. Atrial fibrillation is a type of cardiac arrhythmia that appears as a common finding during medical examinations in humans, large breed dogs and horses. Clinical presentations are of a particular value in racehorses in high performing activities. Atrial fibrillation is characterized by an irregular heart rhythm, secondary to a primary disease or without any sign of comorbidity. The generation and maintenance of Atrial Fibrillation requires a substrate. Some breeds have a genetic predisposition to developing Atrial Fibrillation. Most cases of Atrial Fibrillation are of the paroxysmal type and self-regulate within a few hours to days without the need for treatment. The focus of this study is on the arrhythmic agents that are used for the treatment of Atrial Fibrillation, therefore other arrhythmic agents may not be included, or are included to demonstrate their effect on increasing, inhibiting or decreasing efficacy when used together with medications for the treatment of Atrial Fibrillation. The “working horse” for the pharmacological treatment of Atrial Fibrillation is Quinidine.展开更多
[Objective]The paper was to establish a fluorescence quantitative RT-PCR method for detection of equine arteritis virus(EAV).[Method]Primers and probes were developed for the EAV ORF7 gene sequence,and the reaction sy...[Objective]The paper was to establish a fluorescence quantitative RT-PCR method for detection of equine arteritis virus(EAV).[Method]Primers and probes were developed for the EAV ORF7 gene sequence,and the reaction system was optimized.Standard curves were established,leading to the initial development of the EAV fluorescence quantitative RT-PCR assay.The accuracy,specificity,and sensitivity of this method were subsequently evaluated.[Result]The EAV fluorescence quantitative RT-PCR assay demonstrated optimal performance at an annealing temperature of 61 C,with a final concentration of primer and probe set at 0.6μmol/L.The plasmid standard demonstrated a strong linear correlation with Ct values within the range of 1.6×10^(7)-1.6×10^(2)copies/μL.The equation of the standard curve was determined to be y=-2.68x+32.88,with an R^(2) value of 0.9927.Consequently,the EAV fluorescence quantitative RT-PCR assay was successfully established.The methodology employed was effective in detecting EAV,Theileria equi,equine herpesvirus-1(EHV-1),equine herpesvirus-4(EHV-4),and equine influenza virus(EIV).The findings indicated that the method was specifically capable of detecting EAV,while the other pathogens tested yielded negative results.The method demonstrated a high degree of specificity.It was employed to detect the standard plasmid cRNA synthesized through in vitro transcription following a 10-fold dilution.The results indicated that the minimum detection limit of the method was 1.6×10^(2) copies/μL,and it exhibited high sensitivity.The coefficient of variation,both within and between groups,was maintained at 1.8%,indicating good reproducibility.In this study,the fluorescence quantitative RT-PCR assay developed was utilized alongside the EAV fluorescence quantitative RT-PCR assay established by previous researchers to analyze a total of 234 clinical samples.Both methods yielded a positive detection rate of 14.1%,and the coincidence rate between the two techniques was found to be 100%.[Conclusion]The fluorescence quantitative RT-PCR assay developed in this study offers a novel approach and concept for the prevention and control of equine viral arteritis(EVA).展开更多
Host ANP32 family proteins are crucial for maintaining the activity of influenza RNA polymerase and play an important role in the cross-species transmission of influenza viruses.To date,the molecular properties of equ...Host ANP32 family proteins are crucial for maintaining the activity of influenza RNA polymerase and play an important role in the cross-species transmission of influenza viruses.To date,the molecular properties of equine ANP32(eqANP32)protein are poorly understood,particularly the mechanisms that affect equine influenza virus(EIV)RNA polymerase activity.Here,we found that there are six alternative splicing variants of equine ANP32A(eqANP32A)with different levels of expression.Further studies showed that these six splicing variants of eqANP32A supported the activity of EIV RNA polymerase to varying degrees,with the variant eqANP32A_X2 having the highest expression abundance and exhibiting the highest support of polymerase activity.Sequence analysis demonstrated that the differences in the N-Cap regions of the six splicing variants significantly affected their N-terminal conformation,but did not affect their ability to bind RNA polymerase.We also demonstrated that there is only one transcript of eqANP32B,and that this transcript showed only very low support to the EIV RNA polymerase.This functional defect in eqANP32B is caused by the sequence of the 110–259 amino acids at its Cterminus.Our results indicated that it is the eqANP32A_X2 protein that mainly determines the efficiency of the EIV replication in horses.In conclusion,our study parsed the molecular properties of eqANP32 family proteins and revealed the sequence features of eqANP32A and eqANP32B,suggesting for the first time that the N-Cap region of ANP32A protein also plays an important role in supporting the activity of the influenza virus polymerase.展开更多
The envelope protein （Env） of lentiviruses such as HIV, SIV, FIV and EIAV is larger than that of other retroviruses. The Chinese EIAV attenuated vaccine is based on Env and has helped to successfully control this vi...The envelope protein （Env） of lentiviruses such as HIV, SIV, FIV and EIAV is larger than that of other retroviruses. The Chinese EIAV attenuated vaccine is based on Env and has helped to successfully control this virus, demonstrating that envelope is crucial for vaccine. We compared Env variation of the four kinds of lentiviruses. Phylogenetic analysis showed that the evolutionary relationship of Env between HIV and SIV was the closest and they appeared to descend from a common ancestor, and the relationship of HIV and EIAV was the furthest. EIAV had the shortest Env length and the least number of potential N-linked glyeosylation sites （PNGS） as well as glyeosylation density compared to various immunodefieiency viruses. However, HIV had the longest Env length and the most PNGS. Moreover, the alignment of HIV and SIV showed that PNGS were primarily distributed within extraeellular membrane protein gp120 rather than transmembrane gp41. It implies that the size difference among these viruses is associated with a lentivirus specific function and also the diversity of env. There arc low levels of modification of glycosylation sites of Env and selection of optimal protective epitopes might be useful for development of an effective vaccine against HIV/AIDS.展开更多
Background： The use of equine bone marrow mesenchymal stem cells （BMSC） is a novel method to improve fracture healing in horses. However, additional research is needed to identify optimal culture conditions and to ...Background： The use of equine bone marrow mesenchymal stem cells （BMSC） is a novel method to improve fracture healing in horses. However, additional research is needed to identify optimal culture conditions and to determine the mechanisms involved in regulating BMSC differentiation into osteoblasts. The objectives of the experiments were to determine： 1） if autologous or commercial serum is better for proliferation and differentiation of equine BMSC into osteoblasts, and 2） the expression of key transcription factors during the differentiation of equine BMSC into osteoblasts. Equine BMSC were isolated from the sterna of 3 horses, treated with purchased fetal bovine serum （FBS） or autologous horse serum （HS）, and cell proliferation determined. To induce osteoblast differentiation, cells were incubated with L-ascorbic acid-2-phosphate and glycerol-2-phosphate in the presence or absence of human bone morphogenetic protein2 （BMP2）, dexamethasone （DEX）, or combination of the two. Alkaline phosphatase （ALP） activity, a marker of osteoblast differentiation, was determined by ELISA. Total RNA was isolated from differentiating BMSC between d 0 to 18 to determine expression of runt-reloted tronscrJption foctor2 （Runx2）, osterix （Osx）, and T-box3 （Tbx3）. Data were analyzed by ANOVA. Results： Relative to control, FBS and HS increased cell number （133 ± 5 and 116 ± 5%, respectively; P 〈 0.001） and 5-bromo- 2＇-deoxyuridine （BrdU） incorporation （167 ± 6 and 120 ± 6%, respectively; P 〈 0.001）. Treatment with DEX increased ALP activity compared with control （1,638 ± 38%; P 〈 0.001）. In the absence and presence of Dex, BMP-2 did not alter ALP activity （P 〉 0.8）. Runt-reloted transcription foctor2 expression increased 3-fold （P 〈 0.001） by d 6 of culture. Osterix expression increased 94old （P 〈 0.05） by d 18 of culture. Expression of Tbx3 increased 1.8-fold at d 3 （P 〈 0.01）; however expression was reduced 4-fold at d 18 （P 〈 0.01）. Conclusions： Dexamethasone, but not BMP-2, is required for differentiation of equine BMSC into osteoblasts. In addition, expression of Runx2 and osterix increased and expression of Tbx3 is reduced during differentiation.展开更多
Equine herpesvirus-1 (EHV-1) remains one of the most common viral pathogens affecting horses worldwide presenting as a persistent infection which can establish latency in nerve ganglia (trigeminal ganglion), lymphoid ...Equine herpesvirus-1 (EHV-1) remains one of the most common viral pathogens affecting horses worldwide presenting as a persistent infection which can establish latency in nerve ganglia (trigeminal ganglion), lymphoid tissues of the respiratory tract and peripheral blood lymphocytes. EHV-1 infection induces both humoral and cellular immune responses in horses. Virus neutralising antibody, particularly in the nasopharynx, is to kill free virus shed from infected epithelial cells. Hence this antibody has important functions in reducing virus shedding and spreading infection to cohorts. Cellular immune responses, particularly those carried out by cytotoxic T lymphocyte (CTL), have been shown to be effective in killing virus-infected cells in vitro. This review underlines the state of knowledge regarding immunity to EHV-1 and also its interaction with equine lymphocyte. Finally, the review also includes the importance of the viral immediate early (IE) protein in the pathogenesis of EHV-1. This information can be used as the basis for future research.展开更多
Equine infectious anemia virus(EIAV)is a member of the lentivirus genus in the Retroviridae family and is considered an animal model for HIV/AIDS research.An attenuated EIAV vaccine,which was successfully developed in...Equine infectious anemia virus(EIAV)is a member of the lentivirus genus in the Retroviridae family and is considered an animal model for HIV/AIDS research.An attenuated EIAV vaccine,which was successfully developed in the 1970s by classical serial passage techniques,is the first and only lentivirus vaccine that has been widely used to date.Restriction factors are cellular proteins that provide an early line of defense against viral replication and spread by interfering with various critical steps in the viral replication cycle.However,viruses have evolved specific mechanisms to overcome these host barriers through adaptation.The battle between the viruses and restriction factors is actually a natural part of the viral replication process,which has been well studied in human immunodeficiency virus type 1(HIV-1).EIAV has the simplest genome composition of all lentiviruses,making it an intriguing subject for understanding how the virus employs its limited viral proteins to overcome restriction factors.In this review,we summarize the current literature on the interactions between equine restriction factors and EIAV.The features of equine restriction factors and the mechanisms by which the EIAV counteract the restriction suggest that lentiviruses employ diverse strategies to counteract innate immune restrictions.In addition,we present our insights on whether restriction factors induce alterations in the phenotype of the attenuated EIAV vaccine.展开更多
A simple, rapid, sensitive and reproducible method for enantiomer analysis of methamphetamine, amphetamine, cathinone and methcathinone was developed and validated. The compounds were extracted from equine plasma and ...A simple, rapid, sensitive and reproducible method for enantiomer analysis of methamphetamine, amphetamine, cathinone and methcathinone was developed and validated. The compounds were extracted from equine plasma and urine using a fast liquid-liquid extraction procedure. Only one milliliter plasma and one hundred microliter urine sample is needed for analysis. The extraction procedure had good recovery (>70%) and the matrix effect was negligible. Enantiomer differentiation and confirmation were achieved using liquid chromatography with chiral stationary phase and mass spectrometry detection. The method demonstrated excellent reproducibility with intra-day and inter-day precision of lower than 5%. The lower limits of detection for all of the compounds studied here were at low pg/mL level for both plasma and urine. This is the first report of the analysis of four chiral compounds in equine plasma and urine. Routine application was demonstrated for (S)- and (R)-enantiomer differentiation.展开更多
The aim of this study was to determine the genomic evolutionary pattern of virulent equine infectious anemia virus （EIAV） during persistent infection. The evolutionary dynamics of proviral genomes were examined by c...The aim of this study was to determine the genomic evolutionary pattern of virulent equine infectious anemia virus （EIAV） during persistent infection. The evolutionary dynamics of proviral genomes were examined by challenging an EIAV seronegative equine （pony 1） and three EIAV vaccinated equines （ponies 4, 7, and 8） with the Chinese virulent strain EIAV- L. Ponies 1 and 7 succumbed to disease and were called progressors, while ponies 4 and 8 lacked clinical symptoms and were considered nonprogressors. Sequences spanning the V3, V4, and V5 hyper-variable regions of the EIAV-L envelope gp90 gene were sequenced from each pony as evolutionary markers of the provirus. The proviral genome of the EIAV-L inoculum evolved during persistent infection and displayed different patterns between EIA progressors and nonprogressors. Inoculum-like variants were isolated from nonprogressors during persistent infection, but only from progressors during acute infection. Variant mutations from nonprogressors were dispersed throughout the sequenced region, while those from progressors were predominantly localized to V3. Humoral immunity and virus variant population selection analyses indicated that immune selection was positive in chronically infected progressors and weak in nonprogressors. In-frame stop codons were frequently localized to a defect ＂hot spot＂. The high number of defective variants in nonprogressors may promote disease survival.展开更多
The effects of equine chorionic gonadotropin （eCG） on follicular development and ovulation in cyclic guinea pigs were investigated by histological and immunohistochemical analyses. Three groups of guinea pigs （n=12...The effects of equine chorionic gonadotropin （eCG） on follicular development and ovulation in cyclic guinea pigs were investigated by histological and immunohistochemical analyses. Three groups of guinea pigs （n=12） were administrated subcutaneously with saline, 20 or 50 IU of eCG, respectively, on cyclic Day 12 （Day 1=vaginal openings）. Ovaries were collected at 4 and 8 d after administration （6 animals per group each time）. The eCG administration induced significant and distinct morphological changes in the ovaries, as it promoted the luteinization of granulosa cells, but not follicular development. In addition, proliferating cell nuclear antigen （PCNA） and steroidogenic acute regulatory protein （STAR） were immunolocalized specifically in luteinized follicles. Our experiments together indicate that eCG administration can induce follicular luteinization but not superovulation in guinea pigs. The eCG in cyclic guinea pigs functions similar to that of luteinizing hormone （LH）, but not follicle-stimulating hormone （FSH）.展开更多
Objective:Equine-assisted interventions(EAI)can improve a variety of health problems in older adults and thus promote their well-being.This systematic review aimed to synthesize studies on EAI to understand better the...Objective:Equine-assisted interventions(EAI)can improve a variety of health problems in older adults and thus promote their well-being.This systematic review aimed to synthesize studies on EAI to understand better their effects on the health of older adults.Method:A systematic search guided by the PRISMA 2020 approach was performed on specific databases:Medline(PubMed),EMBASE,PsycINFO,and Cochrane Library.Peer-reviewed articles published in the English language from inception to June 2022 were retrieved.Methodological quality was established using the modified version of the Downs and Black checklist.Results:A total of 244 studies were retrieved,and 13 eligible studies were finally included.Three health domains were investigated:physical(balance,gait,and muscular strength),psychological(quality of life and cognitive assessment),and physiological(hormonal measures,cerebral and muscular activity).Among the eight studies investigating the physical dimension,four studies highlighted a positive effect of EAI on balance,four for gait,and three for strength.Regarding the three studies investigating the psychological dimension,two studies showed a positive effect of EAI on quality of life.Lastly,the four studies investigating the physiological dimensions all demonstrated a positive effect of EAI on hormonal measures and cerebral and muscular activity.Conclusion:Nevertheless,this systematic review provides promising findings regarding the positive effects of EAI on physical,psychological,and physiological health in older adults.Research on EAI should therefore be pursued rigorously to promote this non-pharmacological intervention in an older adult population.展开更多
Equine infectious anemia virus strain L (EIAV-L) is the parental virulent virus of equine infectious anemia donkey leukocyte attenuated vaccine (DLA EIAV). In this study, peripheral blood leukocytes(PBL) were collecte...Equine infectious anemia virus strain L (EIAV-L) is the parental virulent virus of equine infectious anemia donkey leukocyte attenuated vaccine (DLA EIAV). In this study, peripheral blood leukocytes(PBL) were collected from a horse infected with EIAV-L.The PBL DNAs were extracted.The EIAV-L proviral DNA was amplified in four parts covering the entire proviral genomic sequence by polymerase chain reaction (PCR). Each of the four parts was cloned into the plasmid pBluescript SK, and the recombinant plasmids were designated as p2.8, p2.4. p3.1, and p1.2 respectively. After identification with restriction digestion, the inserts within the four plasmids were sequenced. The complete nucleotide sequence of EIAV-L provirus was determined by analyzing each of the four parts and connecting them as a whole. The genome of EIAV-L is 8235 bp in length, and G + C content is 38%. The comparison analysis by the computer software DNASIS showed that the sequence of EIAV-L shares 98.4% and 96.9% identities with that of D-A EIAV and DLA EIAV respectively. The high homology between these strains showed that they were genetically related. The homology between EIAV-L and D-A EIAV is higher than that between EIAV-L and DLA EIAV, and this is consistent with the derivation progress of DLA EIAV. At both ends of EIAV-L provirus, there is an identical long terminal repeat (LTR) sequence of 316bp in length. The LTR consists of U3, R, and U5 regions. The genome of EIAV-L provirus has three long open reading frames(ORF) corresponding to gag, pol and env genes respectively. The gag gene is 1200bp and located at position 613-1912nt. The pol gene is 3402bp and located at position 1708-5109nt. There is a termination codon within the env dividing it into two parts, envl of 699bp (position 5305-6003nt)and env2 of 1827bp (position 6073-7899nt). The provirus has three additional small ORFs: S1, S2 and S3 with sizes of 153bp(position 5113-5265nt), 204bp(position 5279-5482nt)and 402bp(position 7245-7646nt) respectively.展开更多
Selection of proper reference genes (RGs) is an essential step needed for accurate normalization of results from genomic studies. Expression of RGs is regulated by many factors such as species, age, gender, type of ti...Selection of proper reference genes (RGs) is an essential step needed for accurate normalization of results from genomic studies. Expression of RGs is regulated by many factors such as species, age, gender, type of tissue, the presence of disease, and the administration of therapeutic treatment. The aim of the present study was to identify optimal RGs in a set of blood samples collected at different time points (0, 24, 48, 72 h) from horses following administration of extracorporeal shock wave therapy (ESWT). The mRNA expression of twelve RGs: HPRT1, ACTB, HSP90A, SDHA, GUSB, B2M, UBC, NONO, TBP, H6PD, RPL32, GAPDH was determined using real time quantitative polymerase chain reaction (qPCR). An SAS program developed on the algorithm of geNorm, SASqPCR, was used to determine stability of the expression and the number of optimal RGs. The results showed that the range of quantification cycle (Cq) values of the evaluated genes varied between 17 and 26 cycles, and that one optimal RG, ACTB, was sufficient for normalization of gene expression. Results of stability of expression demonstrated that ACTB was the optimal choice for all the samples studied. Notably, in samples collected at 72 h post ESWT, TBP showed a significant change in the expression level, and was not suitable for use as a RG. These results substantiate the importance of validating and selecting an appropriate RG.展开更多
The pathogenesis of equine Osteoarthritis（OA） is more complex, and the disease in the early stage is not easy to be found, therefore, the early diagnosis and treatment are very important. Based on this, this experim...The pathogenesis of equine Osteoarthritis（OA） is more complex, and the disease in the early stage is not easy to be found, therefore, the early diagnosis and treatment are very important. Based on this, this experiment established OA model induced by equine, aimed to study the changes of contents of Matrix Metalloproteinases-3（MMP-3）, Matrix Metalloproteinases-13（MMP-13）, Aggrecanase（ADAMTS-5）, Hyaluronic Acid（HA） and Osteocalcin（OCN） in synovial fluid, and establish rapid diagnostic technique for the equine OA. Thirteen Mongolian equines were used in these induction studies. Equines were randomly divided into two groups： the experimental group contained eight equines and the control group contained five equines. The experimental group was to build the equine osteoarthritis model. The induction was done through Intra-articular（IA） injection of 2 m L Amphotericin-B in equines’ left carpal joints. The equine of the control group was injected into 2 m L physiological saline in equines’ left carpal joints. Synovial fluid was collected every week until the 9th week. The contents of MMP-3, MMP-13, ADAMTS-5, HA and OCN in synovial fluid were evaluated by using ELISA kits. Equine OA model, compared with the control group, starting from the 1st to the 2nd week after induction model, the content of MMP-3, MMP-13, ADAMTS-5, HA and OCN tended to increase, but there was no significant increase, from the 2nd to the 3rd week they significantly increased（p〈0.05） and kept increasing trend until the 9th week. In OA model, MMP-3, MMP-13, ADAMTS-5, HA and OCN showed a rising trend in joint fluid, which would accelerate the cartilage, subchondral bone degradation and metabolism of these proteases increased, and ADAMTS-5 and HA in the early stage increased significantly.展开更多
Equine Infectious Anemia Virus (EIAV) is a retrovirus that establishes a persistent infection in horses and ponies. The virus is in the same lentivirus subgroup that includes human immunodeficiency virus (HIV). The si...Equine Infectious Anemia Virus (EIAV) is a retrovirus that establishes a persistent infection in horses and ponies. The virus is in the same lentivirus subgroup that includes human immunodeficiency virus (HIV). The similarities between these two viruses make the study of the immune response to EIAV relevant to research on HIV. We developed a mathematical model of within-host EIAV infection dynamics that contains both humoral and cell-mediated immune responses. Analysis of the model yields results on thresholds that would be necessary for a combined immune response to successfully control infection. Numerical simulations are presented to illustrate the results. These findings have the potential to lead to immunological control measures for lentiviral infection.展开更多
[ Objective] In order to construct the specific equine disease-free zone of of the Guangzhou Asian Games, to ensure that the equestrian events of Guangzhou Asian Games be smoothly held. [Methods] The species, quantiti...[ Objective] In order to construct the specific equine disease-free zone of of the Guangzhou Asian Games, to ensure that the equestrian events of Guangzhou Asian Games be smoothly held. [Methods] The species, quantities, distribution and seasonal variations of insects and wild animals in related zones were investigated from 2008 to 2010, and the collected samples of the insects and wild animals were tested in laboratory for related equine diseases. [Results] The investigations indicated that there were some mosquitoes, flies, horseflies, punkies, ticks, bats, wild birds and wild bears in equastrianism venue and peripheral regions of disease-free zone of the Guangzhou Asian Games, the laboratory results of Japa- nese encephalitis, vesicular stomatitis, Nipah virus disease, West Nile fever, and Trypanosomiasis evansi, were all negative. According to the in- vestigations and test results, the risk assessments of insect and wild animal vectors in the prevalence of related equine diseases were made to con- firm that the risk was relatively low or very low, and comprehensive prevention and control measures with prevention as major measures and insecti- cides application and environment managements as supplementary means were made on the basis of the risk assessment conclusions. [ Conclu- sions] This research has laid a solid foundation for the successful building of the first specific equine disease-free zone in our country, ensured the smooth holding of the 16th equastdan competition in Guangzhou Asian Games.展开更多
Standard schedules for equine influenza vaccinations have been published in Finland. However, no knowledge about the actual number of vaccinations has been available. Based on previous preliminary studies in slaughter...Standard schedules for equine influenza vaccinations have been published in Finland. However, no knowledge about the actual number of vaccinations has been available. Based on previous preliminary studies in slaughterhouses and among horse owners, it was suspected that vaccination recommendations are not always followed. The aims were to investigate vaccination status against equine influenza among Finnish racehorses in regional competitions. Vaccination status was investigated： （1） by utilizing database vaccination information of show jumping and dressage horses and （2） by carrying out a survey among horse industry experts. A total of 400 horses were selected at the beginning of the Finnish competition season in 2012 and five experts （key persons） were invited to a repeated survey implemented by Delphi techniques. Overall, 50%-100% of horses were properly vaccinated depending on the source available. Thirty-four percent （34%） of the horses had the competition license but no vaccination markings in the database. Differences between types of competition （show jumping and dressage） and between Southern and Northern parts of Finland were found. Vaccinations of show jumping horses were registered more often than vaccinations of dressage horses. Ninety percent （90%） of unvaccinated horses in the database were eight years or older. It can be concluded that equine influenza vaccination registry coverage among racehorses in Finland is incomplete. This poses a risk of misunderstanding, and the growing responsibility of competition organizers.展开更多
Non-steroidal anti-inflammatory drugs (NSAIDs) are classified as Class 4 agents by the Association of Racing Commissioners International and are banned in racehorses during competition in Pennsylvania (PA). To control...Non-steroidal anti-inflammatory drugs (NSAIDs) are classified as Class 4 agents by the Association of Racing Commissioners International and are banned in racehorses during competition in Pennsylvania (PA). To control the abuse of these agents in racehorses competing in PA, a forensic method for screening and confirmation of the presence of these agents is needed. Equine plasma (0.5 mL) was acidified with 75 μL 1M H3PO4 to increase recovery of the analytes by liquid-liquid extraction using methyl tert-butyl ether (MTBE). Extracted analytes were separated by reversed-phase liquid chromatography using a C8 column under gradient condition. All 16 analytes were detected, quantified and confirmed using a triple quadrupole tandem mass spectrometry with selected reaction monitoring (SRM) in both negative and positive electrospray ionization modes. The limit of detection, quantification and confirmation of the analytes were 1.0 - 5.0 ng/mL, 1.0 - 5.0 ng/mL and 1.0 - 20 ng/mL, respectively. The linear dynamic range of quantification was 5.0 - 200 ng/mL. The method is routinely used in anti-doping analysis to control the abuse of NSAIDs in racehorses competing in PA.展开更多
文摘Cardiac arrhythmias are probably more common in horses than in any other domestic animal species where poor performance and exercise intolerance is the most frequent clinical complaint. Atrial fibrillation is a type of cardiac arrhythmia that appears as a common finding during medical examinations in humans, large breed dogs and horses. Clinical presentations are of a particular value in racehorses in high performing activities. Atrial fibrillation is characterized by an irregular heart rhythm, secondary to a primary disease or without any sign of comorbidity. The generation and maintenance of Atrial Fibrillation requires a substrate. Some breeds have a genetic predisposition to developing Atrial Fibrillation. Most cases of Atrial Fibrillation are of the paroxysmal type and self-regulate within a few hours to days without the need for treatment. The focus of this study is on the arrhythmic agents that are used for the treatment of Atrial Fibrillation, therefore other arrhythmic agents may not be included, or are included to demonstrate their effect on increasing, inhibiting or decreasing efficacy when used together with medications for the treatment of Atrial Fibrillation. The “working horse” for the pharmacological treatment of Atrial Fibrillation is Quinidine.
基金Supported by Research Project of General Administration of Customs(2022HK126)Youth Science Foundation(2022D01B08).
文摘[Objective]The paper was to establish a fluorescence quantitative RT-PCR method for detection of equine arteritis virus(EAV).[Method]Primers and probes were developed for the EAV ORF7 gene sequence,and the reaction system was optimized.Standard curves were established,leading to the initial development of the EAV fluorescence quantitative RT-PCR assay.The accuracy,specificity,and sensitivity of this method were subsequently evaluated.[Result]The EAV fluorescence quantitative RT-PCR assay demonstrated optimal performance at an annealing temperature of 61 C,with a final concentration of primer and probe set at 0.6μmol/L.The plasmid standard demonstrated a strong linear correlation with Ct values within the range of 1.6×10^(7)-1.6×10^(2)copies/μL.The equation of the standard curve was determined to be y=-2.68x+32.88,with an R^(2) value of 0.9927.Consequently,the EAV fluorescence quantitative RT-PCR assay was successfully established.The methodology employed was effective in detecting EAV,Theileria equi,equine herpesvirus-1(EHV-1),equine herpesvirus-4(EHV-4),and equine influenza virus(EIV).The findings indicated that the method was specifically capable of detecting EAV,while the other pathogens tested yielded negative results.The method demonstrated a high degree of specificity.It was employed to detect the standard plasmid cRNA synthesized through in vitro transcription following a 10-fold dilution.The results indicated that the minimum detection limit of the method was 1.6×10^(2) copies/μL,and it exhibited high sensitivity.The coefficient of variation,both within and between groups,was maintained at 1.8%,indicating good reproducibility.In this study,the fluorescence quantitative RT-PCR assay developed was utilized alongside the EAV fluorescence quantitative RT-PCR assay established by previous researchers to analyze a total of 234 clinical samples.Both methods yielded a positive detection rate of 14.1%,and the coincidence rate between the two techniques was found to be 100%.[Conclusion]The fluorescence quantitative RT-PCR assay developed in this study offers a novel approach and concept for the prevention and control of equine viral arteritis(EVA).
基金the National Natural Science Foundation of China to HL Zhang(32002275)Natural Science Foundation of Heilongjiang Province of China to HL Zhang(YQ2020C021).
文摘Host ANP32 family proteins are crucial for maintaining the activity of influenza RNA polymerase and play an important role in the cross-species transmission of influenza viruses.To date,the molecular properties of equine ANP32(eqANP32)protein are poorly understood,particularly the mechanisms that affect equine influenza virus(EIV)RNA polymerase activity.Here,we found that there are six alternative splicing variants of equine ANP32A(eqANP32A)with different levels of expression.Further studies showed that these six splicing variants of eqANP32A supported the activity of EIV RNA polymerase to varying degrees,with the variant eqANP32A_X2 having the highest expression abundance and exhibiting the highest support of polymerase activity.Sequence analysis demonstrated that the differences in the N-Cap regions of the six splicing variants significantly affected their N-terminal conformation,but did not affect their ability to bind RNA polymerase.We also demonstrated that there is only one transcript of eqANP32B,and that this transcript showed only very low support to the EIV RNA polymerase.This functional defect in eqANP32B is caused by the sequence of the 110–259 amino acids at its Cterminus.Our results indicated that it is the eqANP32A_X2 protein that mainly determines the efficiency of the EIV replication in horses.In conclusion,our study parsed the molecular properties of eqANP32 family proteins and revealed the sequence features of eqANP32A and eqANP32B,suggesting for the first time that the N-Cap region of ANP32A protein also plays an important role in supporting the activity of the influenza virus polymerase.
基金Natural Science Foundation of China(30970162)Tianjin Municipal Science and Technology Foundation(08ZCGHHZ01800)
文摘The envelope protein （Env） of lentiviruses such as HIV, SIV, FIV and EIAV is larger than that of other retroviruses. The Chinese EIAV attenuated vaccine is based on Env and has helped to successfully control this virus, demonstrating that envelope is crucial for vaccine. We compared Env variation of the four kinds of lentiviruses. Phylogenetic analysis showed that the evolutionary relationship of Env between HIV and SIV was the closest and they appeared to descend from a common ancestor, and the relationship of HIV and EIAV was the furthest. EIAV had the shortest Env length and the least number of potential N-linked glyeosylation sites （PNGS） as well as glyeosylation density compared to various immunodefieiency viruses. However, HIV had the longest Env length and the most PNGS. Moreover, the alignment of HIV and SIV showed that PNGS were primarily distributed within extraeellular membrane protein gp120 rather than transmembrane gp41. It implies that the size difference among these viruses is associated with a lentivirus specific function and also the diversity of env. There arc low levels of modification of glycosylation sites of Env and selection of optimal protective epitopes might be useful for development of an effective vaccine against HIV/AIDS.
基金supported by Storrs Agricultural Experiment Station Hatch Project CONS00844(KEG)
文摘Background： The use of equine bone marrow mesenchymal stem cells （BMSC） is a novel method to improve fracture healing in horses. However, additional research is needed to identify optimal culture conditions and to determine the mechanisms involved in regulating BMSC differentiation into osteoblasts. The objectives of the experiments were to determine： 1） if autologous or commercial serum is better for proliferation and differentiation of equine BMSC into osteoblasts, and 2） the expression of key transcription factors during the differentiation of equine BMSC into osteoblasts. Equine BMSC were isolated from the sterna of 3 horses, treated with purchased fetal bovine serum （FBS） or autologous horse serum （HS）, and cell proliferation determined. To induce osteoblast differentiation, cells were incubated with L-ascorbic acid-2-phosphate and glycerol-2-phosphate in the presence or absence of human bone morphogenetic protein2 （BMP2）, dexamethasone （DEX）, or combination of the two. Alkaline phosphatase （ALP） activity, a marker of osteoblast differentiation, was determined by ELISA. Total RNA was isolated from differentiating BMSC between d 0 to 18 to determine expression of runt-reloted tronscrJption foctor2 （Runx2）, osterix （Osx）, and T-box3 （Tbx3）. Data were analyzed by ANOVA. Results： Relative to control, FBS and HS increased cell number （133 ± 5 and 116 ± 5%, respectively; P 〈 0.001） and 5-bromo- 2＇-deoxyuridine （BrdU） incorporation （167 ± 6 and 120 ± 6%, respectively; P 〈 0.001）. Treatment with DEX increased ALP activity compared with control （1,638 ± 38%; P 〈 0.001）. In the absence and presence of Dex, BMP-2 did not alter ALP activity （P 〉 0.8）. Runt-reloted transcription foctor2 expression increased 3-fold （P 〈 0.001） by d 6 of culture. Osterix expression increased 94old （P 〈 0.05） by d 18 of culture. Expression of Tbx3 increased 1.8-fold at d 3 （P 〈 0.01）; however expression was reduced 4-fold at d 18 （P 〈 0.01）. Conclusions： Dexamethasone, but not BMP-2, is required for differentiation of equine BMSC into osteoblasts. In addition, expression of Runx2 and osterix increased and expression of Tbx3 is reduced during differentiation.
文摘Equine herpesvirus-1 (EHV-1) remains one of the most common viral pathogens affecting horses worldwide presenting as a persistent infection which can establish latency in nerve ganglia (trigeminal ganglion), lymphoid tissues of the respiratory tract and peripheral blood lymphocytes. EHV-1 infection induces both humoral and cellular immune responses in horses. Virus neutralising antibody, particularly in the nasopharynx, is to kill free virus shed from infected epithelial cells. Hence this antibody has important functions in reducing virus shedding and spreading infection to cohorts. Cellular immune responses, particularly those carried out by cytotoxic T lymphocyte (CTL), have been shown to be effective in killing virus-infected cells in vitro. This review underlines the state of knowledge regarding immunity to EHV-1 and also its interaction with equine lymphocyte. Finally, the review also includes the importance of the viral immediate early (IE) protein in the pathogenesis of EHV-1. This information can be used as the basis for future research.
基金supported by grants from the National Natural Science Foundation of China (32172831 and 31672578 to XF.W,32170169 to X.W)the Postdoctoral Scientific Research Developmental Fund of Heilongjiang Province (LBH-Q20059 to XF.W)the Central Public-interest Scientific Institution Basal Research Fund (1610302022007 to XF.W).
文摘Equine infectious anemia virus(EIAV)is a member of the lentivirus genus in the Retroviridae family and is considered an animal model for HIV/AIDS research.An attenuated EIAV vaccine,which was successfully developed in the 1970s by classical serial passage techniques,is the first and only lentivirus vaccine that has been widely used to date.Restriction factors are cellular proteins that provide an early line of defense against viral replication and spread by interfering with various critical steps in the viral replication cycle.However,viruses have evolved specific mechanisms to overcome these host barriers through adaptation.The battle between the viruses and restriction factors is actually a natural part of the viral replication process,which has been well studied in human immunodeficiency virus type 1(HIV-1).EIAV has the simplest genome composition of all lentiviruses,making it an intriguing subject for understanding how the virus employs its limited viral proteins to overcome restriction factors.In this review,we summarize the current literature on the interactions between equine restriction factors and EIAV.The features of equine restriction factors and the mechanisms by which the EIAV counteract the restriction suggest that lentiviruses employ diverse strategies to counteract innate immune restrictions.In addition,we present our insights on whether restriction factors induce alterations in the phenotype of the attenuated EIAV vaccine.
文摘A simple, rapid, sensitive and reproducible method for enantiomer analysis of methamphetamine, amphetamine, cathinone and methcathinone was developed and validated. The compounds were extracted from equine plasma and urine using a fast liquid-liquid extraction procedure. Only one milliliter plasma and one hundred microliter urine sample is needed for analysis. The extraction procedure had good recovery (>70%) and the matrix effect was negligible. Enantiomer differentiation and confirmation were achieved using liquid chromatography with chiral stationary phase and mass spectrometry detection. The method demonstrated excellent reproducibility with intra-day and inter-day precision of lower than 5%. The lower limits of detection for all of the compounds studied here were at low pg/mL level for both plasma and urine. This is the first report of the analysis of four chiral compounds in equine plasma and urine. Routine application was demonstrated for (S)- and (R)-enantiomer differentiation.
基金supported by grants from the National High Technology R&D Program of China (863 Program,2001AA223041)the National Natural Science Foundation of China (30170706)
文摘The aim of this study was to determine the genomic evolutionary pattern of virulent equine infectious anemia virus （EIAV） during persistent infection. The evolutionary dynamics of proviral genomes were examined by challenging an EIAV seronegative equine （pony 1） and three EIAV vaccinated equines （ponies 4, 7, and 8） with the Chinese virulent strain EIAV- L. Ponies 1 and 7 succumbed to disease and were called progressors, while ponies 4 and 8 lacked clinical symptoms and were considered nonprogressors. Sequences spanning the V3, V4, and V5 hyper-variable regions of the EIAV-L envelope gp90 gene were sequenced from each pony as evolutionary markers of the provirus. The proviral genome of the EIAV-L inoculum evolved during persistent infection and displayed different patterns between EIA progressors and nonprogressors. Inoculum-like variants were isolated from nonprogressors during persistent infection, but only from progressors during acute infection. Variant mutations from nonprogressors were dispersed throughout the sequenced region, while those from progressors were predominantly localized to V3. Humoral immunity and virus variant population selection analyses indicated that immune selection was positive in chronically infected progressors and weak in nonprogressors. In-frame stop codons were frequently localized to a defect ＂hot spot＂. The high number of defective variants in nonprogressors may promote disease survival.
基金supported by the National Natural Science Foundation of China(No.31172206)
文摘The effects of equine chorionic gonadotropin （eCG） on follicular development and ovulation in cyclic guinea pigs were investigated by histological and immunohistochemical analyses. Three groups of guinea pigs （n=12） were administrated subcutaneously with saline, 20 or 50 IU of eCG, respectively, on cyclic Day 12 （Day 1=vaginal openings）. Ovaries were collected at 4 and 8 d after administration （6 animals per group each time）. The eCG administration induced significant and distinct morphological changes in the ovaries, as it promoted the luteinization of granulosa cells, but not follicular development. In addition, proliferating cell nuclear antigen （PCNA） and steroidogenic acute regulatory protein （STAR） were immunolocalized specifically in luteinized follicles. Our experiments together indicate that eCG administration can induce follicular luteinization but not superovulation in guinea pigs. The eCG in cyclic guinea pigs functions similar to that of luteinizing hormone （LH）, but not follicle-stimulating hormone （FSH）.
文摘Objective:Equine-assisted interventions(EAI)can improve a variety of health problems in older adults and thus promote their well-being.This systematic review aimed to synthesize studies on EAI to understand better their effects on the health of older adults.Method:A systematic search guided by the PRISMA 2020 approach was performed on specific databases:Medline(PubMed),EMBASE,PsycINFO,and Cochrane Library.Peer-reviewed articles published in the English language from inception to June 2022 were retrieved.Methodological quality was established using the modified version of the Downs and Black checklist.Results:A total of 244 studies were retrieved,and 13 eligible studies were finally included.Three health domains were investigated:physical(balance,gait,and muscular strength),psychological(quality of life and cognitive assessment),and physiological(hormonal measures,cerebral and muscular activity).Among the eight studies investigating the physical dimension,four studies highlighted a positive effect of EAI on balance,four for gait,and three for strength.Regarding the three studies investigating the psychological dimension,two studies showed a positive effect of EAI on quality of life.Lastly,the four studies investigating the physiological dimensions all demonstrated a positive effect of EAI on hormonal measures and cerebral and muscular activity.Conclusion:Nevertheless,this systematic review provides promising findings regarding the positive effects of EAI on physical,psychological,and physiological health in older adults.Research on EAI should therefore be pursued rigorously to promote this non-pharmacological intervention in an older adult population.
基金by National Science Foundation of China(No.39470535).
文摘Equine infectious anemia virus strain L (EIAV-L) is the parental virulent virus of equine infectious anemia donkey leukocyte attenuated vaccine (DLA EIAV). In this study, peripheral blood leukocytes(PBL) were collected from a horse infected with EIAV-L.The PBL DNAs were extracted.The EIAV-L proviral DNA was amplified in four parts covering the entire proviral genomic sequence by polymerase chain reaction (PCR). Each of the four parts was cloned into the plasmid pBluescript SK, and the recombinant plasmids were designated as p2.8, p2.4. p3.1, and p1.2 respectively. After identification with restriction digestion, the inserts within the four plasmids were sequenced. The complete nucleotide sequence of EIAV-L provirus was determined by analyzing each of the four parts and connecting them as a whole. The genome of EIAV-L is 8235 bp in length, and G + C content is 38%. The comparison analysis by the computer software DNASIS showed that the sequence of EIAV-L shares 98.4% and 96.9% identities with that of D-A EIAV and DLA EIAV respectively. The high homology between these strains showed that they were genetically related. The homology between EIAV-L and D-A EIAV is higher than that between EIAV-L and DLA EIAV, and this is consistent with the derivation progress of DLA EIAV. At both ends of EIAV-L provirus, there is an identical long terminal repeat (LTR) sequence of 316bp in length. The LTR consists of U3, R, and U5 regions. The genome of EIAV-L provirus has three long open reading frames(ORF) corresponding to gag, pol and env genes respectively. The gag gene is 1200bp and located at position 613-1912nt. The pol gene is 3402bp and located at position 1708-5109nt. There is a termination codon within the env dividing it into two parts, envl of 699bp (position 5305-6003nt)and env2 of 1827bp (position 6073-7899nt). The provirus has three additional small ORFs: S1, S2 and S3 with sizes of 153bp(position 5113-5265nt), 204bp(position 5279-5482nt)and 402bp(position 7245-7646nt) respectively.
文摘Selection of proper reference genes (RGs) is an essential step needed for accurate normalization of results from genomic studies. Expression of RGs is regulated by many factors such as species, age, gender, type of tissue, the presence of disease, and the administration of therapeutic treatment. The aim of the present study was to identify optimal RGs in a set of blood samples collected at different time points (0, 24, 48, 72 h) from horses following administration of extracorporeal shock wave therapy (ESWT). The mRNA expression of twelve RGs: HPRT1, ACTB, HSP90A, SDHA, GUSB, B2M, UBC, NONO, TBP, H6PD, RPL32, GAPDH was determined using real time quantitative polymerase chain reaction (qPCR). An SAS program developed on the algorithm of geNorm, SASqPCR, was used to determine stability of the expression and the number of optimal RGs. The results showed that the range of quantification cycle (Cq) values of the evaluated genes varied between 17 and 26 cycles, and that one optimal RG, ACTB, was sufficient for normalization of gene expression. Results of stability of expression demonstrated that ACTB was the optimal choice for all the samples studied. Notably, in samples collected at 72 h post ESWT, TBP showed a significant change in the expression level, and was not suitable for use as a RG. These results substantiate the importance of validating and selecting an appropriate RG.
基金Supported by the National Science and Technology Support Program of China(2012BAD46B02-03)
文摘The pathogenesis of equine Osteoarthritis（OA） is more complex, and the disease in the early stage is not easy to be found, therefore, the early diagnosis and treatment are very important. Based on this, this experiment established OA model induced by equine, aimed to study the changes of contents of Matrix Metalloproteinases-3（MMP-3）, Matrix Metalloproteinases-13（MMP-13）, Aggrecanase（ADAMTS-5）, Hyaluronic Acid（HA） and Osteocalcin（OCN） in synovial fluid, and establish rapid diagnostic technique for the equine OA. Thirteen Mongolian equines were used in these induction studies. Equines were randomly divided into two groups： the experimental group contained eight equines and the control group contained five equines. The experimental group was to build the equine osteoarthritis model. The induction was done through Intra-articular（IA） injection of 2 m L Amphotericin-B in equines’ left carpal joints. The equine of the control group was injected into 2 m L physiological saline in equines’ left carpal joints. Synovial fluid was collected every week until the 9th week. The contents of MMP-3, MMP-13, ADAMTS-5, HA and OCN in synovial fluid were evaluated by using ELISA kits. Equine OA model, compared with the control group, starting from the 1st to the 2nd week after induction model, the content of MMP-3, MMP-13, ADAMTS-5, HA and OCN tended to increase, but there was no significant increase, from the 2nd to the 3rd week they significantly increased（p〈0.05） and kept increasing trend until the 9th week. In OA model, MMP-3, MMP-13, ADAMTS-5, HA and OCN showed a rising trend in joint fluid, which would accelerate the cartilage, subchondral bone degradation and metabolism of these proteases increased, and ADAMTS-5 and HA in the early stage increased significantly.
文摘Equine Infectious Anemia Virus (EIAV) is a retrovirus that establishes a persistent infection in horses and ponies. The virus is in the same lentivirus subgroup that includes human immunodeficiency virus (HIV). The similarities between these two viruses make the study of the immune response to EIAV relevant to research on HIV. We developed a mathematical model of within-host EIAV infection dynamics that contains both humoral and cell-mediated immune responses. Analysis of the model yields results on thresholds that would be necessary for a combined immune response to successfully control infection. Numerical simulations are presented to illustrate the results. These findings have the potential to lead to immunological control measures for lentiviral infection.
基金the help and strong support of Departments the Chinese Animal Health and Epidemic Center,Guangdong Rsearch Institute of Entomology,Harbin Veterinary Research Institute of Chinese Academy of Agricultural Sciences,the Agriculture,Fisheries and Conservation Department of Hong Kong and the Hong Kong Jockey Club etc.,during the entomophily and wild animals prevention and control in the disease-free zone of Guangzhou Asian Games
文摘[ Objective] In order to construct the specific equine disease-free zone of of the Guangzhou Asian Games, to ensure that the equestrian events of Guangzhou Asian Games be smoothly held. [Methods] The species, quantities, distribution and seasonal variations of insects and wild animals in related zones were investigated from 2008 to 2010, and the collected samples of the insects and wild animals were tested in laboratory for related equine diseases. [Results] The investigations indicated that there were some mosquitoes, flies, horseflies, punkies, ticks, bats, wild birds and wild bears in equastrianism venue and peripheral regions of disease-free zone of the Guangzhou Asian Games, the laboratory results of Japa- nese encephalitis, vesicular stomatitis, Nipah virus disease, West Nile fever, and Trypanosomiasis evansi, were all negative. According to the in- vestigations and test results, the risk assessments of insect and wild animal vectors in the prevalence of related equine diseases were made to con- firm that the risk was relatively low or very low, and comprehensive prevention and control measures with prevention as major measures and insecti- cides application and environment managements as supplementary means were made on the basis of the risk assessment conclusions. [ Conclu- sions] This research has laid a solid foundation for the successful building of the first specific equine disease-free zone in our country, ensured the smooth holding of the 16th equastdan competition in Guangzhou Asian Games.
文摘Standard schedules for equine influenza vaccinations have been published in Finland. However, no knowledge about the actual number of vaccinations has been available. Based on previous preliminary studies in slaughterhouses and among horse owners, it was suspected that vaccination recommendations are not always followed. The aims were to investigate vaccination status against equine influenza among Finnish racehorses in regional competitions. Vaccination status was investigated： （1） by utilizing database vaccination information of show jumping and dressage horses and （2） by carrying out a survey among horse industry experts. A total of 400 horses were selected at the beginning of the Finnish competition season in 2012 and five experts （key persons） were invited to a repeated survey implemented by Delphi techniques. Overall, 50%-100% of horses were properly vaccinated depending on the source available. Thirty-four percent （34%） of the horses had the competition license but no vaccination markings in the database. Differences between types of competition （show jumping and dressage） and between Southern and Northern parts of Finland were found. Vaccinations of show jumping horses were registered more often than vaccinations of dressage horses. Ninety percent （90%） of unvaccinated horses in the database were eight years or older. It can be concluded that equine influenza vaccination registry coverage among racehorses in Finland is incomplete. This poses a risk of misunderstanding, and the growing responsibility of competition organizers.
文摘Non-steroidal anti-inflammatory drugs (NSAIDs) are classified as Class 4 agents by the Association of Racing Commissioners International and are banned in racehorses during competition in Pennsylvania (PA). To control the abuse of these agents in racehorses competing in PA, a forensic method for screening and confirmation of the presence of these agents is needed. Equine plasma (0.5 mL) was acidified with 75 μL 1M H3PO4 to increase recovery of the analytes by liquid-liquid extraction using methyl tert-butyl ether (MTBE). Extracted analytes were separated by reversed-phase liquid chromatography using a C8 column under gradient condition. All 16 analytes were detected, quantified and confirmed using a triple quadrupole tandem mass spectrometry with selected reaction monitoring (SRM) in both negative and positive electrospray ionization modes. The limit of detection, quantification and confirmation of the analytes were 1.0 - 5.0 ng/mL, 1.0 - 5.0 ng/mL and 1.0 - 20 ng/mL, respectively. The linear dynamic range of quantification was 5.0 - 200 ng/mL. The method is routinely used in anti-doping analysis to control the abuse of NSAIDs in racehorses competing in PA.
