Metagenomic characterization of the equine endometrial microbiome during anestrus.

The equine uterus is highly interrogated during estrus prior to breeding and establishing pregnancy. Many studies in mares have been performed during estrus under the influence of high estrogen concentrations, including the equine estrual microbiome. To date, it is unknown how the uterine microbiome of the mare is influenced by cyclicity; while, the equine vaginal microbiome is stable throughout the estrous cycle. We hypothesized that differences would exist between the equine endometrial microbiome of mares in estrus and anestrus. The aim of this study was two-fold: to characterize the resident endometrial microbiome of healthy mares during anestrus and to compare this with estrus. Double-guarded endometrial swabs were taken from healthy mares during estrus (n = 16) and in the following non-breeding season during anestrus (n = 8). Microbial population was identified using 16S rRNA sequencing. Our results suggest that the equine uterine microbiome in estrus has a low diversity and low richness, while during anestrus, a higher diversity and higher richness were seen compared to estrus. Despite this difference, both the estrus and anestrus endometrial microbiome were dominated by Proteobacteria, Firmicutes, and Bacteroidota. The composition of the microbial community between anestrus and estrus was significantly different. This may be explained by the difference in the composition of the endometrial immune milieu based on the stage of the cycle. Further research investigating the function of the equine endometrial microbiome and dynamics changes within the uterine environment is required.

The Impact of the Composition on the Properties of Simulated Lunar Mare Basalt Fibers.

Lunar mare basalt is recognized as an important in situ resource on the lunar surface. However, the significant compositional variability of lunar mare basalts introduces uncertainties concerning the potential for their use in fabricating fibers and composite materials. This study investigates the impact of different components on the fiber-forming capabilities of mare basalts by simulating the compositions of basalts collected from several well-known lunar missions and then preparing simulated lunar mare basalt fibers. Raman spectroscopy is primarily employed for analysis and characterization, using "peak area normalization" to explore the impact of compositional fluctuations in the simulated lunar mare basalts on the glass network structure. The findings indicate that an increase in the Fe content raises the likelihood of basalt fibers crystallizing. Additionally, Fe3+ is shown to substitute for Si and Al in constructing bridging oxygen bonds in the network structure, albeit reducing the overall polymerization of the network. Meanwhile, Fe2+ acts as a network modifier to enhance the mechanical properties of the fibers.

[Influence of lactoferrin and enzymatiс hydrolysates of cow's and mare's milk proteins on anaphylactic sensitivity and cytokine profile of rats].

The development of innovative ingredients of specialized formula for children with intolerance to cow's milk proteins requires accounting the influence of the protein component on the allergic sensitivity. The aim of the research was to study the effect of lactoferrin (LF) from cow colostrum, cow's milk protein hydrolysate (CMPH) and mare's milk protein hydrolysate (MMPH) on the severity of the systemic anaphylaxis reaction, the levels of specific IgG antibodies and cytokines in rats parenterally sensitized with ovalbumin (OVA). Material and methods. The experiment was carried out on 4 groups of 26 male Wistar rats, which were sensitized intraperitoneally with chicken egg OVA and a systemic anaphylaxis reaction was induced on the day 29 by intravenous administration of a challenge dose of the antigen (6 mg per kg body weight). LF, CMPH and MMPH were introduced into the diet in doses of 1.4-2 g/kg body weight per day (on an average 1.59±0.04, 1.53±0.05 and 1.48±0.05 g per kg body weight respectively). The content of IgG antibodies in the blood serum was determined by an indirect ELISA; the levels of cytokines IL-1α, IL-1ß, IL-2, IL-4, IL-5, IL-6, IL-10, IL-12(p70), IL-13, GM-CSF, IFN-γ and TNF-α were detected by multiplex immunoassay. Results. Dietary LF did not have a significant effect on the severity of active anaphylactic shock (AAS), concentrations of antibodies and cytokines in the blood of animals. As a result of CMPH consumption, there were no significant changes in AAS severity and IgG antibodies to OVA but significant increase in TNF-α level was observed as well as a significant decrease in IL-1α (p<0.05). In animals receiving MMPH, there were also no significant changes in the severity of AAS, but a 1.9-fold decrease in the level of IgG antibodies to OVA (p<0.001) was noticed along with a significant increase in IL-12(p70) (p<0.05) and IL-10 (p<0.10) level. Conclusion. Cosumption of LF by sensitized rats didn't significantly affect their anaphylactic sensitivity and cytokine profile, while CMPH intake induced some signs of pro-inflammatory processes. Consumption of MMPH was accompanied by the formation of an anti-inflammatory cytokine profile, which corresponds to a decrease in the intensity of the humoral immune response to the model allergen. Differences in the effects of two hydrolisates, which have similar degrees of hydrolysis, may be associated with the specific composition of glycopeptides formed during the enzymatic cleavage of milk protein produced by these two species of dairy animals.

Effect of pregnancy on isolation efficiency and in vitro proliferation of equine peripheral-blood derived mesenchymal stromal cells.

Mesenchymal stromal cells (MSCs) have regenerative and immunomodulatory potential and may be used to treat injured tissues. Pregnancy has been associated with increased MSCs in the peripheral circulation in multiple species, but to date, there are no reports on this matter in horses. This study aimed to evaluate the effect of pregnancy on isolation efficiency and proliferation capacity of equine MSCs derived from the peripheral blood (PB) of mares. Venous blood samples were collected at the 11th month of gestation and 1 month after delivery from clinically healthy Arabian mares that presented normal pregnancies. Blood samples were processed for in vitro cellular culture and hormonal and metabolic profiles. MSCs were isolated and characterized by trilineage differentiation potential, immunophenotyping, analyzed by gene sequencing and proliferation assays. The isolation of peripheral blood mononuclear cells (PBMCs) of pregnant mares were associated with higher isolation efficiency and proliferative capacity of MSCs derived from peripheral blood (PB-MSCs) recovered pre-partum than those isolated post-partum. Although fetal gender, parity, 5α-reduced pregnanes, insulin, and cortisol were shown to affect cellular proliferation, individual factors and the small population studied must be considered. This study suggests that PB-MSCs from pregnant mares could be a valuable alternative source of MSCs for therapeutic purposes.

Isolation of 60 strains from fermented milk of mares and donkeys in Algeria and identification by 16S rRNA sequencing of lactobacilli: Assessment of probiotic skills of important strains and aromatic productivity power.

Background and Aim: Donkey and mare milk have high nutritional and functional values, but their lactic acid bacteria (LAB) content remains poorly studied and undervalued in the Algerian dairy industry. This study aimed to isolate and select LAB strains that produce antimicrobial substances during fermentation and to characterize the probiotic profiles of each extracted strain to indicate their potential for antioxidant and proteolytic activity. Materials and Methods: This study focuses on isolating and identifying lactic acid bacterial strains from 10 Equid-fermented milk samples collected in two regions of El Bayed Wilaya (Algeria). Identification of LAB strains was obtained by 16S rRNA sequencing. The probiotic properties of important strains and their aromatic productivity power are assessed. To evaluate their antibacterial activity against Listeria monocytogenes, Staphylococcus aureus, Chryseobacterium joostei, Pseudomonas aeruginosa, and Escherichia coli, we selected 21 strains. Different induction methods have been used to amplify the antibacterial effects against these pathogenic strains. Results: Among a total of 60 identified strains, 31 had a probiotic profile, and most were catalase-negative. Aromatic productivity power was observed in eight strains: Lactiplantibacillus plantarum, Lactobacillus casei, Lactobacillus paracasei, Weissella confusa, Weissella cibaria, Leuconostoc mesenteroides, Leuconostoc lactis, and Lactobacillus sp1. Conclusion: Our results provide insight into the considerable diversity of LAB present in fermented donkey and mare milk. To meet the expectations of the Algerian dairy industry, it is important that the probiotic skills of the nine selected strains are met. In addition, a significant number of these strains may have important probiotic activity and biotechnological potential.

Use of cytobrush for bacteriological and cytological diagnosis of endometritis in mares.

Background and Aim: A combined microbial and cytological examination of uterine samples is the main diagnostic method for endometritis in mares. This study aimed to describe a procedure for using the same uterine cytobrush (CB) for both bacteriological and cytological evaluation. Material and Methods: The procedure consists of rolling the CB onto a sterilized glass slide immediately after collection and before the transfer into a sterile saline solution. In Experiment 1, a comparison between bacteriological results of the cotton swab (CS) and CB or pellet was made in 10 mares; in Experiment 2, bacteriological and cytological results were compared between different processing methods of CB in 28 mares; in other 6 mares, a CB was processed for cytology only, to investigate the reasons for the low cellularity of the pellet. Results: The agreement between culture results from the CB and CS was evaluated, and a comparison between the cytological data obtained by different processing methods of CB was performed. The perfect agreement between the CB and CS microbiological results was found. The described procedure enables useful diagnostic smears for cytology. Moreover, the seeding of both the tip of CB and the saline solution used for the transport produced accurate bacteriological results. Conclusion: The protocol described in this study for the use of CB for both cytological and bacteriological analysis could be used for the diagnosis of endometritis. To maximize diagnostic sample quality, cytology slides must be prepared with meticulous care in the field to preserve cellular integrity and minimize artifacts.

Characterization of Typical Behaviors of Mares in the Opening Phase of Parturition-Influence of Parity and Dystocia.

The identification of typical behaviors in stage I of parturition, the opening phase, can be used to improve birth monitoring in the mare. Therefore, this study aimed to comprehensively analyze mare behavior during the opening phase. Real-time recordings of 66 births involving 56 warmblood mares were analyzed using camera footage. Behaviors such as increased locomotor activity, pawing with front hooves, lifting the tail, rolling, kicking with the hind legs toward the abdomen, and looking at the abdomen increased significantly (p < 0.001) in the four hours preceding parturition. Within the last hour of the observation period, a statistically significant change was observed for the duration of lying in the sternal and lateral position (p < 0.001). Significant correlations were observed between parity and the total number of repetitions of lying in the sternal position (p < 0.05). Furthermore, the birth process influenced the repetitions of lying in the lateral position (p < 0.05). These findings indicate distinct behavioral patterns during the opening phase of parturition, which were evident across the observed mares. Nonetheless, notable individual differences were also identified among the mares.

Effect of age on androgens pattern in cyclic mares.

Androgens are produced in both sexes. In females produced by the adrenal gland and the ovaries they play a crucial role in regulating ovarian function, estrogen synthesis and follicular growth. Age leads to a reduction in androgen concentrations, although, at present, these mechanisms are not elucidated in mares. The objective of this study was to evaluate the concentrations of testosterone (T), androstenedione (A4) and dehydroepiandrosterone (DHEA) in mares of different ages. Blood samples were drawn from seventy cyclic Spanish Purebred mares belonging to five age groups: 3-5 years, 6-9 years, 10-13 years, 14-16 years and > 16 years. The concentrations of T, A4 and DHEA were determined by EIA, validated specifically for horses. Mares aged 3-5, 6-9 and 10-13 years had higher T concentrations (P < 0.05) than mares aged >16 years, and mares aged 6-9 years had also higher concentrations than those 14-16 years old (P < 0.05). A4 concentrations were lower (P < 0.05) in mares >16 years old when compared with those of other age groups. DHEA concentrations were lower (P < 0.05) in mares 14-16 years and > 16 years old when compared with those of other age groups. DHEA was positively correlated with T (r = 0.61; P < 0.05) and A4 (r = 0.51; P < 0.05). Age induces reduction in androgens' synthesis in physiologically cyclic Spanish Purebred mares. These physiological variations must be duly considered for a correct and objective interpretation of the analytical data.

Evaluation of peritoneal l-lactate concentration in horses in the early post-partum period.

BACKGROUND: Peritoneal fluid lactate concentration is an important diagnostic tool in horses with abdominal pain. Information on peritoneal lactate concentrations is lacking following parturition in the mare. OBJECTIVES: To compare blood and peritoneal lactate concentrations in a population of mares within 36 h post-partum, report a normal reference range and identify any impact of retained foetal membranes (RFMs). METHODS: This is a retrospective study evaluating healthy mares from which blood and peritoneal samples had been obtained within 36 h of parturition. Exclusion criteria included signs of abdominal pain within this period. Data was interrogated for normality using a Shapiro-Wilk test. Wilcoxon signed-rank test and Bland-Altman analysis were used to compare blood and peritoneal lactate concentrations. Linear regression was used to compare age and breed data with peritoneal lactate concentrations. Significance was defined as p < 0.05. RESULTS: Forty mares met the inclusion criteria. Mean age was 12.6 ± 4.1 years, and most mares were multiparous (65%). Peritoneal lactate ((1.2 (IQR = 0.9-1.6) mmol/L) was increased compared to blood lactate concentration (0.7 (IQR = 0-1.1)mmol/L; p < 0.001). Plasma total protein (TP) concentrations were 68 (IQR = 64-74) g/L and peritoneal protein concentrations 8 (IQR = 4-9.7) g/L. Six mares developed RFM. The median fold-increase in peritoneal lactate concentration compared to blood lactate concentration was 0.9 (IQR: 0.01-1.7; range: 0-2.5). The reference range for peritoneal fluid lactate concentration was 0-2.5 mmol/L. CONCLUSION: Peritoneal lactate concentrations in healthy post-partum mares remained within the normal reference range and were not influenced by RFM or parturition. Increased peritoneal lactate in this group warrants further investigation.

Identification of pasteurized mare milk and powder adulteration with bovine milk using quantitative proteomics and metabolomics approaches.

Adulteration in dairy products presents food safety challenges, driven by economic factors. Processing may change specific biomarkers, thus affecting their effectiveness in detection. In this study, proteomics and metabolomics approaches were to investigate the detection of bovine milk (BM) constituents adulteration in pasteurized mare milk (PMM) and mare milk powder (MMP). Several bovine proteins and metabolites were identified, with their abundances in PMM and MMP increasing upon addition of BM. Proteins like osteopontin (OPN) and serotransferrin (TF) detected adulteration down to 1 % in PMM, whereas these proteins in MMP were utilized to identify 10 % adulteration. Biotin and N6-Me-adenosine were effective in detecting adulteration in PMM as low as 10 % and 1 % respectively, while in MMP, their detection limits extend down to 0.1 %. These findings offer insights for authenticating mare milk products and underscore the influence of processing methods on biomarker levels, stressing the need to consider these effects in milk product authentication.

In vitro effects of different ozone preparations on microorganisms responsible for endometritis in the mare.

Infectious endometritis is considered one of the major causes of infertility and it can affect up to 60% of barren mares. It is characterized by the presence of one or more microorganisms in the reproductive tract and it is treated with the administration of antibiotics, ecbolic agents and uterine lavages. Ozone, thanks to its antimicrobial properties that are based on its high oxidative potential, could represent an effective alternative treatment for endometritis. The aim of this study was to test in vitro the bactericidal and fungicidal properties of different ozone formulations, either as gas (experiment 1) or dissolved in two liquid matrices (experiment 2), specifically distilled water or oil (Neozone 4000, Cosmoproject, Parma, Italy), onto 6 different species of microorganisms isolated from mares with clinical endometritis, namely Escherichia coli, Staphylococcus aureus, Streptococcus equi subsp. Zooepidemicus, Pseudomonas aeruginosa, Klebsiella pneumoniae and Candida albicans. In the first experiment, 3 clinical antibiotic-resistant strains per each species were exposed to different conditions: to O2O3 gas mixtures (15 and 40 µg/ml for 1, 3 and 5 min), to 100 % O2 or left untreated. The results showed a reduction of the microbial count of over 99,9% for every pathogen, time and concentration of O2O3 gas mixtures tested. Furthermore, gaseous ozone showed both a time-dependant effect (5 vs 3 vs 1 min of exposure) and a concentration-dependant effect (40 vs 15 µg/ml) at 1 and 3 min, while after 5 min no differences were observed. In the second experiment, minimum inhibitory concentration (MIC), and minimum bactericidal/fungicidal concentration (MBC, MFC) of ozonated distilled water and ozonated oil were evaluated. Ozonated oil showed a bactericidal/fungicidal activity against all the strains tested (MIC range 12.5-25 % v/v, MBC/MFC range 12.5-50 % v/v) while ozonated distilled water didn't show an observable antimicrobial effect, discouraging its use as an antimicrobial agent for the treatment of endometritis. The results of this in vitro study indicate that both gaseous ozone and ozonated oil exerted remarkable antimicrobial activities and are promising alternative treatments for infectious endometritis, even when caused by antibiotic-resistant bacteria, and encourage further experiments in an effort to scale down or even prevent the use of antibiotics in equine reproduction.

Characterizing the relationship between gonadotropin releasing hormone (GnRH), kisspeptin, and RFamide related peptide 3 (RFRP-3) neurons in the equine hypothalamus across the estrous cycle and in the anovulatory seasons.

To understand better the role that kisspeptin plays in regulating seasonal and estrous cycle changes in the mare, this study investigated the number, location and interactions between GnRH, kisspeptin and RFRP-3 neurons in the equine hypothalamus. Hypothalami were collected from mares during the non-breeding season, vernal transition and various stages of the breeding season. Fluorescent immunohistochemistry was used to label the neuropeptides of interest. GnRH cells were observed primarily in the arcuate nucleus (ARC), while very few labeled cells were identified in the pre-optic area (POA). Kisspeptin cells were identified primarily in the ARC, with a small number of cells observed dorsal to the ARC, surrounding the third ventricle (3V). The mean number of kisspeptin cells varied between animals and typically showed no pattern associated with season or stage of estrous cycle, but a seasonal difference was identified in the ARC population. Small numbers of RFRP-3 cells were observed in the ARC, ventromedial hypothalamus (VMH) and dorsomedial hypothalamus (DMH). The mean number of RFRP-3 cells appeared higher in pre-ovulatory animals compared to all other stages. The percentage of GnRH cell bodies with kisspeptin appositions did not change with season or stage of estrous cycle. The percentage of kisspeptin cells receiving inputs from RFRP-3 fibers did not vary with season or stage of estrous cycle. These interactions suggest the possibility of the presence of an ultra-short loop feedback system between these three peptides. The changes in RFRP-3 neurons suggest the possibility of a role in the regulation of reproduction in the horse, but it is unlikely to be as a gonadotropin inhibitory factor.

Follicular Fluid-Derived Extracellular Vesicles Influence on In Vitro Maturation of Equine Oocyte: Impact on Cumulus Cell Viability, Expansion and Transcriptome.

Cumulus cell (CC) expansion is pivotal for oocyte maturation, during which CCs release factors that initiate paracrine signaling within the follicular fluid (FF). The FF is abundant in extracellular vesicles (EVs) that facilitate intercellular communication. Although bovine and murine EVs can control cumulus expansion, these effects have not been observed in equines. This study aimed to assess the impact of FF-derived EVs (ffEVs) on equine CC expansion, viability, and transcriptome. Cumulus-oocyte complexes (COCs) that underwent in vitro maturation (IVM) in the presence (200 µg protein/mL) or absence (control) of ffEVs were assessed for cumulus expansion and viability. CCs were isolated after 12 h of IVM, followed by RNA extraction, cDNA library generation, and subsequent transcriptome analysis using next-generation sequencing. Confocal microscopy images illustrated the internalization of labeled ffEVs by CCs. Supplementation with ffEVs significantly enhanced cumulus expansion in both compacted (Cp, p < 0.0001) and expanded (Ex, p < 0.05) COCs, while viability increased in Cp groups (p < 0.01), but decreased in Ex groups (p < 0.05), compared to the controls. Although transcriptome analysis revealed a subtle effect on CC RNA profiles, differentially expressed genes encompassed processes (e.g., MAPK and Wnt signaling) potentially crucial for cumulus properties and, consequently, oocyte maturation.

Prevalence of Microbial Isolates Cultured from Endometrial Swab Samples Collected from United Kingdom Thoroughbred Mares from 2014 to 2020.

Determining whether endometrial microbial isolates are pathogens, contaminants, or even part of the "normal" microbiome is extremely complex, particularly given the absence of "gold standard" tests for endometritis. Population-level benchmarking and temporal monitoring can provide novel insights and a wider context to improve understanding. This study aimed to (i) estimate the prevalence of endometrial isolates from swabs of Thoroughbred broodmares in Newmarket, UK between 2014 and 2020; and (ii) evaluate the effects of year, mare age, and cytology findings on isolate prevalence. Generalised linear mixed models with a logit link, both null models and models using year of sampling, mare age, or cytology findings as predictors, were fitted to estimate isolate prevalence. Over the 7-year period, data were available from 18,996 endometrial-swab samples from 6050 mares on 290 premises. The overall isolate prevalence was 35.5% (95% confidence interval (CI) 33.0-37.9), and this varied significantly between years. The most prevalent isolates were ß-hemolytic Streptococcus (17.9; 95% CI: 17-19) and E. coli (10.3%; 95% CI: 9.0-11.6). Isolate prevalence increased with mare age except for E. coli isolates, and with increasing category of cytology findings except for α-hemolytic Streptococcus isolates. The results provide novel estimates of isolate prevalence and highlight knowledge gaps around potential complexities in the interpretation of findings.

Fermented Mare Milk and Its Microorganisms for Human Consumption and Health.

Mare milk is consumed by approximatively 30 million people in the world. In countries in Asia and East Europe, mare milk is mainly consumed as source of fermented products, called koumiss, airag or chigee, alcoholic beverages obtained by means of a culture of bacteria and lactose-fermenting yeasts. Recent research concerning mare milk and its derivatives deals mainly with their potential employment for human health. Studies about the isolation and characterization of Lactobacillus spp. and yeasts from koumiss have been aimed at assessing the potential functional properties of these micro-organisms and to find their employment for the industrial processing of mare milk. The aim of this literature review is to summarize recent research about microorganisms in fermented mare milk products and their potential functional properties.

A case of non-mosaic X trisomy (65,XXX) in a Thoroughbred mare confirmed by cytogenetic and molecular analysis.

A 9-year-old Thoroughbred mare with normal external genitalia and regular oestrus symptoms was gynecologically examined prior to insemination. This primary examination revealed the presence of a hypoplastic uterus and the lack of normal ovaries, and the mare was therefore subjected to more detailed diagnostics, including endocrinological, genetic, and clinical tests. Diagnostic imaging with the use of ultrasonography and endoscopy confirmed the underdevelopment of internal genitalia. Analysis of circulating sex hormones revealed very low concentrations of progesterone and oestradiol. Finally, cytogenetic analysis showed the presence of non-mosaic X trisomy (65,XXX), an aneuploidy of sex chromosomes that is rarely detected in horses. This finding was also confirmed by molecular methods, including highly sensitive droplet digital PCR (ddPCR) and microsatellite markers genotyping. Our study reveals the need for gynaecological and genetic evaluation of broodmares, even if their phenotype (including developed external genitalia and oestrus symptoms) shows no signs of potential abnormalities.

Success rates and factors influencing pregnancy outcome after 464 transvaginal ultrasound-guided twin reductions in the mare.

BACKGROUND: Transvaginal ultrasound-guided aspiration (TUA) is used for post-fixation twin reduction in mares. However, there is limited information regarding factors that influence pregnancy outcome after TUA. OBJECTIVES: To evaluate the effect of day of gestation on which TUA is performed, aspiration volume, puncture of the conceptus, medication administered before and after TUA, embryo location, mare age and parity and operator experience on pregnancy and foaling rates after TUA. STUDY DESIGN: Retrospective case series. METHODS: Data were collected from case records of 464 TUAs performed by 14 operators in 422 mares diagnosed pregnant with dizygotic twins in two different facilities between 2010 and 2019. Pregnancy status was determined by ultrasonography at 5-7 days and 3-4 weeks after the TUA was performed. Subsequent pregnancy and foaling results were obtained by follow-up communication. The effects of mare, gestation- and TUA-related variables on pregnancy and foaling rates were analysed by the chi-square-test for homogeneity and Fisher's exact test and logistic regression. RESULTS: TUA was performed between 21 and 82 days of gestation in unilaterally (267/359 [74.4%]) and bilaterally fixed (92/359 [25.6%]) twin pregnancies. A singleton pregnancy (218/381 [57.2%]), persistent twin pregnancy (60/381 [15.8%]), or the loss of both conceptuses (103/381 [27%]) was confirmed 5-7 days after TUA was performed. At 3-4 weeks post TUA 50.3% (163/324) of mares were diagnosed with a single viable pregnancy and 40.1% (127/317) went on to deliver a live single foal. TUA performed early in gestation (D 25-35) resulted in the birth of a live singleton foal in 49.3% (74/150) of mares. MAIN LIMITATIONS: Missing retrospective data despite extensive follow-up. CONCLUSION: This is the first large scale study to demonstrate that acceptable pregnancy and foaling rates can be achieved in mares diagnosed with twins when TUA is performed early in gestation (<40 days).

Development of a fluorometric Cuboni test for the semi-quantitative measurement of urinary estrogen levels and pregnancy detection in mares.

The aim of the present study was to develop a semi-quantitative urine pregnancy test for mares based on the Cuboni reaction and to verify the reliability of this test. The urine specimens were hydrolyzed by heating in the presence of hydrochloric acid. The resulting free estrogens were extracted from the urine matrix using toluene. Sulfuric acid was added to the toluene extract and the mixture was heated again. The lower layer in the test tube containing sulfuric acid was used for fluorescence measurements with excitation at 355 nm and measurement at 535 nm. The fluorometric Cuboni test revealed that the fluorescence counts in urine samples collected after the second trimester of gestation were significantly higher than those obtained from barren mares. The levels of estrogens, including equilin, estrone and estardiol-17ß exhibited a dose-dependent increase in fluorescence counts, whereas other steroids, such as progesterone, testosterone, and cortisol, did not affect fluorescence. Heat treatment of urine samples with hydrochloric acid significantly increased the fluorescence counts in those collected after the second trimester of gestation compared to non-pregnant samples, implying the presence of large amounts of conjugated estrogens in pregnant mare urine. Fluorescence counts in urine samples obtained during pregnancy showed a positive relationship with estrone concentrations as measured by enzyme immunoassay. The results of the present study showed that the fluorometric Cuboni test facilitates urine fluorescence counts depending on the urinary estrogen content and is capable of discriminating between pregnancy and non-pregnancy states beyond the second trimester of gestation in mares.

The Effect of Supplemental Lighting during the Late Gestation Period on Post-Partum Mechanical Properties of Mare and Foal Guard Hair.

This study investigates Thoroughbred mares exposed or not to supplemental blue light at the end of the gestation. Sixty mares and their 60 foals were selected for the investigation. Guard hair samples were collected from the shoulder just after (within 12 h) the parturition or birth. The foals of the light-treated mares developed significantly (p < 0.05) shorter hair than those of the control mares. A general effect of light treatment on basal hair diameter thinning could be demonstrated (p < 0.005). The maximum force of hair samples of light-treated mares and foals (0.098 and 0.085 N, respectively) was significantly lower than that of the control (0.272 and 0.178 N, respectively). The tensile strength (82.2 N/mm2) of the foal hair samples of the light-treated mares was significantly lower than that of the control foals (121.6 N/mm2). Although no significant difference was found in the elongation (ΔL), the hair of the control animals (mares and foals together) was more elastic than that of the treated animals (335 vs. 262 µm). In conclusion, the supplemental blue light treatment of the pregnant mares has a decreasing effect on both mares and their foals on the mechanical properties of the hair, making it shorter, thinner, and weaker.

Analysis of the Influence of Lactiplantibacillus plantarum and Lacticaseibacillus rhamnosus Strains on Changes in the Hexachlorobenzene Content in Fermented Mare Milk during Refrigerated Storage.

(1) Background: Hexachlorobenzene (HCB) is a persistent organic pollutant that is possibly carcinogenic to humans. It is still found in the environment, humans and animals, and in foods, including milk and dairy products; (2) Methods: The influence of the probiotic cultures Lacticaseibacillus rhamnosus LCR and Lactiplantibacillus plantarum subsp. plantarum LP on the possibility of effecting the biodegradation of HCB in dairy products fermented from mare milk was investigated, taking into account the product storage time (maximum 21 days). HCB content was determined using the GC/MS method; (3) Results: A strong negative Pearson correlation (p < 0.05) was found between HCB concentration and the refrigeration storage time of the fermented beverages. The highest HCB reduction was observed in milk fermented with both Lacticaseibacillus rhamnosus LCR and Lactiplantibacillus plantarum subsp. plantarum LP (78.77%), while the lowest was noted when only Lactiplantibacillus plantarum subsp. plantarum LP was used (73.79%); (4) Conclusions: This pilot study confirmed that probiotics commonly used to give products health-promoting properties can also contribute to reducing the content of undesirable substances, and the bacterial cultures used might provide an alternative method for reducing HCB residues in fermented drinks.