Horse Serum Research Articles

REFERENCE VALUES OF BLOOD BIOCHEMICAL PARAMETERS FOR THE ZHABE TYPE KAZAKH HORSE IN THE SOUTHERN REGION OF THE REPUBLIC OF KAZAKHSTAN

The reference values for biochemical parameters in blood serum of the zhabe type Kazakh horse are presented in this article. To obtain reference biochemical parameters, whole blood samples of 50 clinically healthy adult zhabe-type horses of both sexes and raised on the free pasture of the "Kalka" farm in Zhambyl region, Shu district, Baluan Sholak, were examined on an automatic biochemical analyzer A15 (Biosystems, Spain). Biochemical results of zhabe type horses were close to the established normal indicators. The levels of enzymes of the hepatobiliary system (aspartate amintransferase, alkaline phosphatase), triglyceride content in horses of the zhabe type were 3-5 times higher than normal. Lower concentrations of albumin and zinc were found (22.22±3.67 g/l and 4.86±1.49 mmol/l, respectively), as well as creatinine concentrations within the lower normal values (77.84±14.19 mmol/L). Therefore, reference intervals of biochemical blood parameters (albumins, total protein, urea, uric acid, cholesterol, creatinine, bilirubin, triglycerides, glucose, calcium, phosphorus, magnesium, iron, zinc, alkaline phosphatase, alanine amintransferase and aspartate amintransferase) were determined for the zhabe type horse in the conditions of the Southern region of the Republic of Kazakhstan, typical for the normal course of metabolic processes in the body.

One-step immunoassay based on filtration for detection of food poisoning-related bacteria

A one-step immunoassay based on filtration was presented, which used microbeads for target analyte detection and filters with appropriate pore sizes to distinguish the complexity of target analyte and microbeads. For effective bacterial detection, the microbead size and the filter's pore size must be optimized. The optimal concentrations of the enzyme (urease) and antibody were determined at the maximum absorbance change, that is, the maximum pH change. The pH change was measured using a field-effect transistor (FET). The correlation between pH change and threshold voltage was estimated to be 21.7 mV/pH, and the correlation between pH change and the source-drain current was estimated to be −379 nA/pH. For the one-step immunoassay, antibodies against target bacteria were isolated from horse serum by filtration, and these antibodies were estimated to have a sufficiently high specificity to overcome cross-reactivity among five types of food poisoning-related bacteria: Escherichia coli O157, Salmonella typhimurium, Listeria monocytogenes, Bacillus cereus, and Staphylococcus aureus. Finally, the FET-based one-step immunoassay was demonstrated for five types of food poisoning-related bacteria in human serum.

Protein microarray allergen profiling in bronchoalveolar lavage fluid and serum of horses with asthma

The diagnostic value of allergen-specific immunoglobulin E (IgE) in horses with asthma is uncertain. A recently developed protein microarray detected abnormally high latex-specific IgE concentrations in the serum of horses with severe asthma. The main objective was to characterize the IgE profiles of asthmatic horses in Switzerland using a protein microarray platform in serum and bronchoalveolar lavage fluid (BALF). The secondary objective was to determine whether serological and BALF allergen-specific IgE concentrations correlated. Forty-four asthmatic and 39 control horses ≥5 years of age. This prospective cross-sectional study investigated the sensitization profiles of horses with asthma compared with environmentally matched healthy controls. Both serum and BALF were analyzed using the protein microarray. Partial least square-discriminant analysis (PLS-DA) was used to identify and rank the importance of the allergens for class detection (ie, asthma vs control), with a variable influence on the projection (VIP) >1 considered significant. The allergens that best discriminated (VIP >1) asthmatic horses from controls were proteins derived from fungi (Aspergillus fumigatus), insects (Culicoides spp.), and latex (Hevea brasiliensis). The serological model predictive ability was markedly inferior (area under the curve [AUC] 0.585, 95% confidence interval [CI]: 0.454-0.747) to that of the BALF (AUC 0.751, 95% CI: 0.582-0.866). The two models shared nine allergens, of which eight showed significant weak to moderate correlations. The concentrations of several allergen-specific IgE were higher in asthmatic horses. The protein microarray performed better on BALF than serum for detection of asthma. Serological IgE concentrations do not closely correlate with BALF concentrations and should be interpreted with caution.

Genetic characterization of three recently discovered parvoviruses circulating in equines in China.

The family Parvoviridae comprises many major viral pathogens that can infect humans and multiple other species, causing severe diseases. However, knowledge of parvoviruses that infect equids is limited. In the present study, we found that three equine parvoviruses (EqPVs), namely, equine parvovirus-hepatitis (EqPV-H), equine parvovirus-cerebrospinal fluid (EqPV-CSF) and equine copivirus (EqCoPV) cocirculated among horses in China. We examined the prevalence of these three EqPVs in 225 horse serum samples in China and found EqPV-H, EqPV-CSF and EqCoPV viremia in 7.6% (17/225), 2.7% (6/225) and 2.2% of samples (5/225), respectively. We also obtained the complete genomes of one EqPV-H strain, six EqPV-CSF strains and one EqCoPV strain. After phylogenetic analysis of the EqPVs, we found that EqPV-CSF and EqCoPV may have evolved from the same ancestor. The EqPV-CSF strains (E111 and A27) and EqCoPV strain (F124) were genetically similar to foreign strains, but the EqPV-CSF strains (B48, E96, C61 and F146) comprised unique clades. This study determined the prevalence of three EqPVs in Chinese horses and analyzed the genetic characteristics of EqPVs prevalent strains in Chinese horse herds. Our data provide a theoretical basis for follow-up research on the prevention and control of EqPVs.

Sensitivity and specificity for African horse sickness antibodies detection using monovalent and polyvalent vaccine antigen-based dot blotting.

The immune responses of animals infected with African horse sickness (AHS) virus are determined by enzyme-linked immunosorbent assay (ELISA), complement fixation, and virus neutralization test. During the outbreaks of AHS in Thailand, the immune response after vaccination has been monitored using commercial test kits such as blocking ELISA, which are expensive imported products unavailable commercially in Thailand. This study aimed to assess the sensitivity and specificity of anti-AHS virus antibodies using dot blotting based on monovalent and polyvalent strains of live attenuated AHS vaccine. A total of 186 horse sera, namely, 93 AHS-unvaccinated samples and 93 AHS-vaccinated samples, were used in this study. All sera underwent antibodies detection using commercial blocking ELISA and in-house dot blotting based on monovalent and polyvalent strains of live attenuated AHS vaccine. The numbers of true positive, false positive, true negative, and false negative results in the dot blotting were compared with those in blocking ELISA and the sensitivity and specificity of dot blotting were assessed. For the monovalent antigen, there were 78, 19, 74, and 15 true positive, false positive, true negative, and false negative results, respectively, while for the polyvalent antigen, the corresponding numbers were 84, 34, 58, and 9. Meanwhile, the diagnostic sensitivity and specificity for monovalent antigen were 83.87% and 79.57%, respectively, but 90.32% and 62.37% for polyvalent antigen. Dot blotting for AHS antibodies detection using vaccine antigen showed high sensitivity and rather a high specificity compared with the findings with the commercial ELISA test kit. In countries where commercial ELISA test kits are not available and when the size of a serum sample is small, dot blotting could become a good alternative test given its advantages, including its simplicity, rapidity, and convenience. To the best of our knowledge, these findings are the first report on the use of dot blotting for detecting AHS antibodies in horses. In conclusion, monovalent antigen-based dot blotting could be used as a reliable alternative serodiagnostic test for monitoring AHS humoral immune response, especially in vaccinated horses.

Chicken Serum과 Horse Serum이 닭 근육위성세포의 증식과 분화에 미치는 영향

For efficient production of cultured meat, chicken breast muscle cells and chicken leg muscle cells were proliferated and differentiated using medium with various serums, and proliferation and differentiation ability were analyzed. Chicken breast muscle cells and chicken leg muscle cells were cultured with serum-free medium (NC), medium with 20% FBS (C), medium with 10% chicken serum (CS) and 5% horse serum (HS) (T1), medium with 10% CS (T2), medium with 5% HS (T3). Using the immunofluorescence staining, Pax7 and nuclei were stained in chicken breast muscle cells and chicken leg muscle cells to confirm that they were muscle satellite cells. As a result of cell counting, both cells cultured in T1 medium proliferated the most. Chicken breast muscle cells and chicken leg muscle cells were subcultured from passage 1 to passage 6 using T1 medium. In both cells, viability and number of live cells decreased with increasing passage, but chicken leg muscle cells had higher viability and number of live cells than chicken breast muscle cells in all passages. Cells were differentiated using differentiation medium containing 10% chicken serum and 5% horse serum for each passage. It was confirmed that the differentiation ability of chicken leg muscle cells was more maintained as passage increased than chicken breast muscle cells. Therefore, it is considered that the results of chicken cells will become the basic data for the development of cultured meat.

Changes in haematological and biochemical parameters in blood serum of horses during exposition to workload stress

Health state of animals undergoing experimental procedures is an important topic nowadays, as even the small changes can influence the outcome of entire outcomes. Main aim of the study was to evaluate the influence of horse training on variety of blood parameters including mineral profile, energy profile, hepatic profile and haematology. In the experiment, the studied group of horses underwent training programme which consisted of transportation, lounging, riding, jumping, racing, treadmill training and shoeing. Blood samples were collected and later evaluated at the beginning, in the middle and at the end of 1-year lasting process. Our results show multiple significant changes in blood parameters, including changes to multiple minerals, such as Ca, K, Na as well as significant changes of total proteins, urea and certain hepatic profile parameters. Haematology results have also been affected in individual sample collections. Based on results of our study we can state that there have been changes to the internal milieu of the horse but also that there have not been any visible changes of the health status of the animals over the duration of the experiment.

A Novel Ex Vivo Drug Assay for Assessing the Transmission-Blocking Activity of Compounds on Field-Isolated Plasmodium falciparum Gametocytes.

The discovery and development of transmission-blocking therapies challenge malaria elimination and necessitate standard and reproducible bioassays to measure the blocking properties of antimalarial drugs and candidate compounds. Most of the current bioassays evaluating the transmission-blocking activity of compounds rely on laboratory-adapted Plasmodium strains. Transmission-blocking data from clinical gametocyte isolates could help select novel transmission-blocking candidates for further development. Using freshly collected Plasmodium falciparum gametocytes from asymptomatic individuals, we first optimized ex vivo culture conditions to improve gametocyte viability and infectiousness by testing several culture parameters. We next pre-exposed ex vivo field-isolated gametocytes to chloroquine, dihydroartemisinin, primaquine, KDU691, GNF179, and oryzalin for 48 h prior to direct membrane feeding. We measured the activity of the drug on the ability of gametocytes to resume the sexual life cycle in Anopheles after drug exposure. Using 57 blood samples collected from Malian volunteers aged 6 to 15 years, we demonstrate that the infectivity of freshly collected field gametocytes can be preserved and improved ex vivo in a culture medium supplemented with 10% horse serum at 4% hematocrit for 48 h. Moreover, our optimized drug assay displays the weak transmission-blocking activity of chloroquine and dihydroartemisinin, while primaquine and oryzalin exhibited a transmission-blocking activity of ~50% at 1 μM. KDU691 and GNF179 both interrupted Plasmodium transmission at 1 μM and 5 nM, respectively. This new approach, if implemented, has the potential to accelerate the screening of compounds with transmission-blocking activity.

The relationship between the intensity of Gasterophilus intestinalis larvae infection and the serum and salivary humoral immune response in horses

Infection with Gasterophilus intestinalis (botfly) larvae often occurs in horses. The aim of the study was to isolate the larvae of G. intestinalis and evaluate the serum and salivary humoral immune response using self-developed ELISA in G. intestinalis infected horses. Blood serum or saliva samples were taken from 125 infected horses and 54 uninfected slaughtered horses. The antigens from G. intestinalis larvae were used for development of ELISA in order to evaluate the intensity of G. intestinalis IgG, IgM, and IgA antibody reactivity in the serum or saliva of naturally infected horses and horses without larvae in the gastrointestinal tract (control group). Serum antibodies against second and third larvae’s stadium antigens reacted significantly more intensively in infected than in healthy horses in IgG (p ≤ 0.001; p ≤ 0.05, respectively) and IgA (p ≤ 0.05;p ≤ 0.001, respectively) classes. Salivary IgG and IgA specific’s antibody reactivity was significantly higher in horses with moderate (p ≤ 0.01) and severe infection (p ≤ 0.001) compared to the healthy horses. The determination of the G. intestinalis IgG and IgA antibody activity in saliva and serum may be used for detecting horses moderately and severely infested with larvae.

Intramuscular injection of a mixture of COVID-19 peptide vaccine and tetanus vaccine in horse induced neutralizing antibodies against authentic virus of SARS-CoV-2 Delta variant

Peptide vaccine is not effective due to its low immunogenicity. To improve the efficacy of peptide vaccine against COVID-19, a novel method was developed by mixing a COVID-19 peptide vaccine with a tetanus vaccine. In this study, intramuscular injection of a mixture of COVID-19 peptide vaccine and tetanus vaccine twice, i.e., first dose on day 0 and second dose on day 21, induced neutralizing antibodies against authentic virus of SARS-CoV-2 Delta variant in a horse. Horse serum of day 35, i.e., two weeks after the second dose, neutralized authentic virus of SARS-CoV-2 Delta variant, equal to half effectiveness of human serum from vaccinees of Moderna COVID-19 vaccine. However, neither horse serum nor human serum neutralized Omicron variant authentic virus. No side effects were observed after each dose. This study indicates intramuscular injection of a mixture of COVID-19 peptide vaccine and tetanus vaccine may work in humans to improve peptide vaccine efficacy against SARS-CoV-2.

A colorimetric assay and MCR-ALS analysis of the peroxidase-like activity of poly (N-phenylglycine) functionalized with polyethylene glycol (PNPG-PEG) nanozyme for the determination of dopamine

This report describes, for the first time, the coupling of UV–visible spectroscopy with multivariate curve resolution-alternative least-squares (MCR-ALS) algorithm to study peroxidase-like catalytic reaction of polyethylene glycol-functionalized poly (N-phenyl glycine) (PNPG-PEG) as an efficient and intrinsic peroxidase mimic activity (PMA) class of conducting organic polymer for selective detection of dopamine (DA) in the PNPG-PEG + TMB + H2O2 reaction system. PNPG-PEG was produced by means of a chemical route using ammonium persulphate (APS) as an oxidant agent of N-phenyl glycine monomer. The chemical composition, morphology, and thermal behavior of PNPG-PEG were examined by various instrumental techniques. PNPG-PEG exhibited significant peroxidase-mimic activity to catalyze the oxidation 3,3′,5,5′- tetramethylbenzidine (TMB) substrate to oxidized TMB (oxTMB). The qualitative and quantitative determination of the oxidized TMB can easily be detected by the naked-eye and the recorded UV–vis absorbance spectra at 652 nm, respectively. Owing to the superior peroxidase-mimic activity of PNPG-PEG, the colorimetric detection of dopamine was successfully achieved at pH 4.0. Under optimal conditions, acceptable linear dependency was recorded in the concentration range of 5.1–125.0 μM, with a limit of detection (LOD) and limit of quantification (LOQ) equal to 4.6 μM and 13.8 μM (S/N = 3), respectively. Furthermore, this colorimetric assay was successfully used for quantitative analysis of dopamine in fetal bovine serum (FBS) and horse serum (HS) samples with recoveries in the range of 97–105% and 100–122%, respectively. After resolving the bilinear data matrix using MCR-ALS, three chemical components were found for different concentrations and pure spectral profiles. Based on the resolved profiles, the presence of free, slightly penetrated, and majorly penetrated TMB molecules entering the polymeric structure can be easily detected using MCR-ALS as an available statical method without any complex separation instruments. This peroxidase mimetic nanozyme as a visual, simple, low-cost, sensitive, and reproducible colorimetric platform can provide great potential applications in the monitoring and diagnosis of dopamine-related diseases.

Expression patterns and correlation analyses of muscle-specific genes in the process of sheep myoblast differentiation.

The purpose of this study was to establish a system for the isolation, culture, and differentiation of sheep myoblasts, and to explore the expression patterns as well as mutual relationships of muscle-specific genes. Sheep fetal myoblasts (SFMs) were isolated by two-step enzymatic digestion, purified by differential adhesion and identified using immunofluorescence techniques. Two percent horse serum was used to induce differentiation in SFMs. Real-time quantitative and Western blot analyses were respectively used to detect the mRNA and protein expressions of muscle-specific genes including MyoD, MyoG, Myf5, Myf6, PAX3, PAX7, myomaker, desmin, MYH1, MYH2, MYH4, MYH7, and MSTN during the differentiation of SFMs. Finally, the correlation between muscle-specific genes was analyzed by the Pearson correlation coefficient method. The results showed that the isolated and purified SFMs could form myotubes after the induction for differentiation. The marker factors including MyoD, MyoG, myomaker, desmin, and MyHC were positively stained in SFMs. The mRNA expressions of MyoD, MyoG, and myomaker increased and then decreased, while Myf5, PAX3, and PAX7 decreased; Myf6, desmin, MYH1, MYH2, MYH4, and MYH7 increased; and MSTN fluctuated up and down during the differentiation of SFMs. The expression patterns of protein were basically consistent with those of mRNA except MSTN. There existed significant or highly significant correlations at mRNA or protein level among some genes. Some transcription factor proteins (MyoD, Myf5, Myf6, PAX3, PAX7) showed significant or highly significant correlations with the mRNA level of some other genes and/or themselves. In conclusion, SFMs with good myogenic differentiation ability were successfully isolated, and the expression patterns and correlations of muscle-specific genes during SFM differentiation were revealed, which laid an important foundation for elucidating the gene regulation mechanism of sheep myogenesis.

PREVALENCE OF Blastocystis sp. IN CATTLE, GOAT AND SHEEP REARED BY DIFFERENT FARM MANAGEMENT SYSTEMS IN PAHANG, MALAYSIA

Blastocystis sp. is a familiar parasite in the gastrointestinal tract causing infection in humans and animals. The purpose of this study is to evaluate the Blastocystis sp. prevalence in three sorts of livestock; cattle, goats, and sheep in Pahang, Malaysia, which are reared under two management systems; intensive and semi-intensive farm management system. About 92, 96, and 65 cattle, goat, and sheep fecal samples respectively were collected from different farms around Kuantan, Bera, and Pekan. The fecal samples were cultured in Jones’ medium supplemented with 10% heat-inactivated horse serum and incubated at 37 ºC for 2 weeks, then observed under light microscopy daily. The total prevalence of Blastocystis sp. was 29.34% in cattle (27/92), 29.16% in goats (28/96), and 43.07% in sheep (28/65). Supported the results of this study, Blastocystis sp. prevalence was higher in sheep and livestock reared by a semi-intensive farm management system (44.38%). However, further study could be done for Blastocystis sp. subtypes identification to determine its genetic diversity. Notwithstanding, this study has provided additional knowledge on the prevalence of each livestock reared in farms around Pahang that serve as important information in understanding host-parasite relationships, besides determining the best farm management system to be applied by farmers.

The transcriptome expression levels related to ovulation induction and acupuncture protection therapy in rats through gene microarray

ABSTRACT This study aimed to explore the potential molecular mechanisms of acupuncture in the adjuvant treatment of infertility. A rat model of ovulation induction was constructed using horse serum gonadotropin and human chorionic gonadotropin, and then acupuncture was used to treat the model rats at Guanyuan and Sanyinjiao points. Hematoxylin eosin (HE) staining, the endometrial thickness, and enlargement of uterine cavity were determined to evaluate the effects of acupuncture on the uterus of ovulation induction rats. Meanwhile, the uterus tissues were sent for gene microarray analyses. Acupuncture enhanced the endometrial receptivity of ovulation induction rats. Gene microarray showed that 189 overlapped differential expressed genes (DEGs) were identified, and these overlapped DEGs were divided into four clusters by Mfuzz algorithm. Afterwards, interaction networks containing 842 interaction pairs were established, and the DEGs in the interaction networks were significantly enriched in 20 BP and 10 KEGG pathways, including wound healing, response to glucocorticoid, TNF signaling pathway, terpenoid backbone biosynthesis, and glutathione metabolism. By calculating topological parameters, Anxa5, Casp8, Tgm2, Frk, Mmp7, Timp1, Hmgcr, Cth, Serpinal, and Abcbla were the important hub genes of the interaction networks. Our findings revealed the changes at transcriptome levels related to ovulation induction and acupuncture protection therapy.

PSIV-A-12 Effects of Treating Primary Bovine Satellite Cells Cultures Inducted to Differentiate with Steroid Hormones, Zinc or Manganese on Creatine Kinase Activity

Abstract Anabolic implants containing estradiol (E2) and trenbolone acetate (TBA) promote skeletal muscle growth by increasing protein synthesis of primary bovine satellite cells (BSC). Recent data indicate skeletal muscle growth induced by anabolic implants may be positively augmented by trace mineral (TM) supplementation. Currently, the effects that E2, TBA or TM have on differentiation of primary BSC cultures is unknown. The objective of this study was to determine whether BSC differentiation is altered when cultures are treated with E2, TBA, zinc (Zn) or manganese (Mn). Primary BSC were grown to 80% confluency and treated (n=2) with 3% horse serum (CON), 10 nM TBA, 10 nM E2, 10 nM TBA and 10 nM E2, or 0.05, 0.10 or 0.25 μM of Mn, or 10, 20 or 40 μM of Zn. Creatine kinase activity was measured at 0, 24 and 72 h post-treatment to assess BSC differentiation. Creatine kinase is a muscle specific enzyme that is upregulated during satellite cell differentiation. A series of contrasts were constructed to compare CON and treated cultures at each time point. Creatine kinase activity increased (P< 0.05) over time indicating all cultures were differentiating. There was no difference (P >0.10) between the CON and any of the treatments at either 0 or 24 h post-differentiation. Treatment with 10 nM TBA increased (P=0.03) creatine kinase activity at 72 h compared to CON cultures. However, treatment with Zn, Mn, E2 or E2 and TBA did not have an effect (P >0.15) on BSC differentiation as measured by creatine kinase activity. This preliminary data indicates TBA might increase BSC differentiation, but further experiments are needed to validate this finding. Further, additional research analyzing the impacts of Zn and Mn in the presence of different steroid hormones is warranted to better understand the relationship that exists between steroid hormones and TM.

P466 Isolation of Cryptococcus neoformans and other yeast from pigeon droppings in Khartoum state, Sudan

Poster session 3, September 23, 2022, 12:30 PM - 1:30 PMObjectivesThe aim of this study was to investigate the presence of Cryptococcus neoformans and other yeast species in pigeon droppings in Khartoum state, Sudan.MethodsSample collection A total of 120 samples were collected from pigeon droppings from Khartoum state. Pigeon droppings were collected using sterile wooden spatulas and placed directly in clean plastic bags.Processing of sampleApproximately 2 g of pigeon droppings were added to 10 ml of sterile saline. The samples were allowed to stand for 20 min with frequent vortex.Isolation and Identification of yeast isolatesA loopful of supernatant fluid from prepared samples was taken and streaked onto Sabouraud's dextrose agar media supplemented with 0.05 mg/ml chloramphenicol incubated at 37°C for 1-2 days. All yeast isolates were identified by direct microscopic examination using the lacto-phenol-cotton-blue stain India ink stain and Gram's stain. Further confirmatory tests were conducted using Corn Meal Agar (CMA) with tween 80, germ tube production test using horse serum, and urease test. Confirmation of identification was done using API 20C AUX and API ID 32C AUX (bioMérieux®, Madrid, Spain).Results Cryptococcus neoformans is the most common species isolated from pigeon droppings as shown in Table 1. Different Candida species have been isolated Figure 1.Isolated yeastNumber Cryptococcus neoformans 42 Candida spp34 Cryptococcus albidus 5 Stephanoscus ciferrii 4 Rhodotorula glutinis 4 Rhodotorula mucilaginosa 3 Geotrichum capitatum 2 Zygosaccharomyces spp1ConclusionThe present study concluded that there is a potential role of pigeons as a reservoir for C. neoformans and other zoonotic yeasts in the environment that can affect humans and animals.

Infection of young foals with Equine Parvovirus-Hepatitis following a fatal non-biologic case of Theiler’s disease

Theiler’s disease (TD) is a (sub-)acute hepatitis in adult horses and one of the most common causes of acute hepatic failure. Recent findings indicate that equine parvovirus hepatitis (EqPV-H) likely causes TD and that its transmission occurs via iatrogenic and/or natural routes. Following the death of an EqPV-H positive mare with TD, close-contact mares and foals in the same paddock were monitored to evaluate if there was any evidence of EqPV-H. For this purpose, the serum of close contact horses was examined 6 and 42 days after the mare’s death for the presence of EqPV-H DNA and changes in liver-associated serum biochemical parameters. The foals had higher EqPV-H viral loads than the mares. Apart from the mare that was euthanized, none of the horses included in this study showed signs of severe disease and nor did they have particularly elevated liver enzymes. Nucleotide sequence analysis revealed no major differences between the viral DNA detected in the serum of the dead mare and any of the in-contact horses. In conclusion, our data confirmed previous findings that horizontal transmission of EqPV-H may occur through close contact between horses.

Rethinking common solvents in butyrylcholinesterase activity assays

• The effect of solvents on enzyme activity is quite diverse. Even two enzymes, which are members of the same family, may have different responses to the same solvent. • Acetone has the highest potential to inhibit the butyrylcholinesterase enzyme. • Butyrylcholinesterase is slightly inhibited by ethanol and methanol, making these solvents comparatively preferable in activity measurement assays. • Owing to their inhibitory potentials, the final concentrations of common solvents in the assay medium must be considered carefully. Cholinesterases are a group of enzymes that prevent the message of the neurotransmitter acetylcholine from being transmitted in an undesirable manner. The primary classes of this family are acetylcholinesterase (AChE) and butyrylcholinesterase (BChE). In neurodegenerative disorders, including Alzheimer’s disease (AD) and Parkinson’s disease, the levels of these enzymes are altered. The level of AChE declines in Alzheimer’s disease, while the level of BChE increases or remains unchanged. There have been a number of licenced medications developed to combat the negative health impacts of these disorders. However, synthetic medications have a number of unfavorable side effects. The search for alternative BChE inhibitors in the treatment of AD is also on the rise. Some of the compounds examined are not soluble in water and must be dissolved in organic solvents. However, these organic solvents may have a negative or detrimental influence on enzymes. This study examined the inhibitory effects of acetone, dimethyl sulfoxide (DMSO), acetonitrile, ethanol, and methanol on horse serum BChE (hsBChE). Competitive inhibition was observed with acetone, DMSO and ethanol, non-competitive inhibition was observed with methanol, and acetonitrile exhibited linear mixed-type competitive inhibition. Acetone, DMSO, acetonitrile, ethanol, and methanol had IC 50 values of 707 mM, 1719 mM, 1848 mM, 7741 mM, and 12199 mM, respectively, and K i values of 186 ± 15 mM; 529 ± 35 mM, 725 ± 67 mM, 3281 ± 29 mM, and 22991 ± 6535 mM, respectively. These IC 50 and K i values show that methanol is the best solvent, with almost minimal interaction with hsBChE.

Infrared spectroscopy of serum fails to identify early biomarker changes in an equine model of traumatic osteoarthritis

Objective: to determine the accuracy of infrared (IR)-based serum biomarker profiling to differentiate horses with early inflammatory changes associated with a traumatically induced model of equine carpal osteoarthritis (OA) from controls.Method: unilateral carpal OA was induced in 9 of 17 healthy Thoroughbred fillies, while the remainder served as sham operated controls. Serum samples were obtained before induction of OA (Day 0) and weekly thereafter until Day 63 from both groups. Films of dried serum were created, and IR absorbance spectra acquired. Following pre-processing, partial least squares discriminant analysis (PLSDA) and principal component analysis (PCA) were used to assess group and time differences and generate predictive models for wavenumber ranges 1300-1800 ​cm−1 and 2600-3700 ​cm−1.Results: the overall correct classification rate when classifying samples by group (OA or Sham) was 52.7% (s.d. ​= ​12.8%), while it was 94.0% (s.d. ​= ​1.4%) by sampling Day. The correct classification results by group-sampling Day combinations with pre-intervention serum (Day 0) was 50.5% (s.d. ​= ​21.7%).Conclusion: with the current approach IR spectroscopic analysis could not differentiate serum of horses with induced carpal OA from that of controls. The high classification rate obtained by Day of sampling may reflect the effect of exercise on the biomarker profile. A longer study period (advanced disease) or naturally occurring disease may provide further information on the suitability of this technique in horses.

Copper deposition in Wilson's disease causes male fertility decline by impairing reproductive hormone release through inducing apoptosis and inhibiting ERK signal in hypothalamic-pituitary of mice.

Wilson’s disease (WD) is an autosomal recessive disorder of copper metabolism characterized by liver and central nervous system dysfunction. Considerable evidence suggests that infertility is also very common in male patients with WD, but the exact molecular mechanisms involved remain unknown. In order to further investigate the pathological changes in the hypothalamic-pituitary-testicular (HPT) axis and its mechanisms, mice were divided into the normal control group (NC), WD model TX mice group (WD), dimercaptosuccinic acid–treated TX mice group (DMSA), and pregnant horse serum gonadotropin–treated TX mice group (PMSG). The copper content and morphology of hypothalamus and pituitary tissues, the ultrastructure and apoptosis of hypothalamus neurons and pituitary gonadotropin cells, the serum levels of reproductive hormones, and the pregnancy rate and litter size of the female mice were studied. The expression of apoptosis-related proteins and the phosphorylation of extracellular regulatory protein kinase (ERK) 1/2 in the hypothalamus and pituitary were detected. The results showed that the copper content was significantly increased in the WD group, and the histopathological morphology and ultrastructure of the hypothalamus and pituitary were damaged. The levels of the gonadotropin-releasing hormone, the follicle-stimulating hormone, the luteinizing hormone, and testosterone were significantly decreased. The apoptosis rate in the hypothalamus and pituitary was significantly increased. The expressions of proapoptotic proteins Bax and Caspase-3 were significantly increased, the expression of the anti-apoptotic protein Bcl-2 was significantly decreased, and the phosphorylation level of ERK1/2 was significantly decreased. Fertility is significantly reduced. After DMSA intervention, the hypothalamus tissue copper content decreased, the hypothalamus and pituitary tissue morphology and ultrastructure were improved, cell apoptosis was alleviated, the expression of Bax and Caspase-3 was significantly decreased, the expression of Bcl-2 was significantly increased, and the reproductive hormone level, phosphorylation level, and fertility were increased. Fertility was preserved after treatment with PMSG in male TX mice. These results suggest that copper deposition in WD causes male fertility decline by impairing reproductive neuroendocrine hormone release through inducing apoptosis and inhibiting the ERK signal in the hypothalamic–pituitary region. This study can also provide reference for the damage of copper pollution to the male reproductive system.