Preclinical modeling of metabolic syndrome to study the pleiotropic effects of novel antidiabetic therapy independent of obesity.

Cardiovascular-kidney-metabolic health reflects the interactions between metabolic risk factors, chronic kidney disease, and the cardiovascular system. A growing body of literature suggests that metabolic syndrome (MetS) in individuals of normal weight is associated with a high prevalence of cardiovascular diseases and an increased mortality. The aim of this study was to establish a non-invasive preclinical model of MetS in support of future research focusing on the effects of novel antidiabetic therapies beyond glucose reduction, independent of obesity. Eighteen healthy adult Beagle dogs were fed an isocaloric Western diet (WD) for ten weeks. Biospecimens were collected at baseline (BAS1) and after ten weeks of WD feeding (BAS2) for measurement of blood pressure (BP), serum chemistry, lipoprotein profiling, blood glucose, glucagon, insulin secretion, NT-proBNP, angiotensins, oxidative stress biomarkers, serum, urine, and fecal metabolomics. Differences between BAS1 and BAS2 were analyzed using non-parametric Wilcoxon signed-rank testing. The isocaloric WD model induced significant variations in several markers of MetS, including elevated BP, increased glucose concentrations, and reduced HDL-cholesterol. It also caused an increase in circulating NT-proBNP levels, a decrease in serum bicarbonate, and significant changes in general metabolism, lipids, and biogenic amines. Short-term, isocaloric feeding with a WD in dogs replicated key biological features of MetS while also causing low-grade metabolic acidosis and elevating natriuretic peptides. These findings support the use of the WD canine model for studying the metabolic effects of new antidiabetic therapies independent of obesity.

Effects of the oral administration of glycosaminoglycans with or without native type II collagen on the articular cartilage transcriptome in an osteoarthritic-induced rabbit model.

BACKGROUND: In a previous study, the 84-day administration of glycosaminoglycans (GAGs), with or without native collagen type II (NC), in an osteoarthritis (OA)-induced rabbit model slowed down OA progression, improved several micro- and macroscopic parameters and magnetic resonance imaging (MRI) biomarkers in cartilage, and increased hyaluronic acid levels in synovial fluid. To elucidate the potential underlying mechanisms, a transcriptomics approach was conducted using medial femoral condyle and trochlea samples. RESULTS: The administration of chondroitin sulfate (CS), glucosamine hydrochloride (GlHCl), and hyaluronic acid (HA), with (CGH-NC) or without (CGH) NC, strongly modulated several genes involved in chondrocyte extracellular matrix (ECM) remodeling and homeostasis when compared to non-treated rabbits (CTR group). Notably, both treatments shared the main mechanism of action, which was related to ECM modulation through the down-regulation of genes encoding proteolytic enzymes, such as ADAM metallopeptidase with thrombospondin type 1 motif, 9 (Adamts9), and the overexpression of genes with a relevant role in the synthesis of ECM components, such as aggrecan (Acan) in both CGH-NC and CGH groups, and fibronectin 1 (Fn1) and collagen type II, alpha 1 (Col2A1) in the CGH group. Furthermore, there was a significant modulation at the gene expression level of the mTOR signaling pathway, which is associated with the regulation of the synthesis of ECM proteolytic enzymes, only in CGH-NC-supplemented rabbits. This modulation could account for the better outcomes concerning the microscopic and macroscopic evaluations reported in these animals. CONCLUSIONS: In conclusion, the expression of key genes involved in chondrocyte ECM remodeling and homeostasis was significantly modulated in rabbits in response to both CGH and CGH-NC treatments, which would partly explain the mechanisms by which these therapies exert beneficial effects against OA.

Paraoxonase I Activity and Its Relationship with Nutrition in Amyotrophic Lateral Sclerosis.

Background: Amyotrophic lateral sclerosis (ALS) is characterized by progressive motor neuron degeneration, with oxidative stress playing a key role. Paraoxonase 1 (PON1) is an antioxidant enzyme that may influence ALS progression. This study aimed to establish a predictive model for the influence of PON1 activity on functionality in ALS patients and explore its relationship with nutrition. Methods: In this observational cross-sectional study, 70 ALS patients underwent assessments of PON1 activity, lipid profile, functional capacity, respiratory function, and heart rate variability. A structural equation model was developed to determine the relationships between variables. Nutritional intake was analyzed in 65 patients. Results: The predictive model showed that PON1 activity and LDL levels positively influenced functionality, both directly and indirectly through respiratory capacity. Heart rate variability moderately predicted functionality independently. HDL levels were not significantly associated with functionality. Weak to moderate correlations were found between PON1 activity and intake of certain nutrients, with positive associations for monounsaturated fats and vitamin D, and negative associations for carbohydrates, proteins, and some micronutrients. Conclusions: PON1 activity appears to play an important role in ALS patient functionality, both directly and through effects on respiratory capacity. However, its relationship with nutritional intake was not strongly evident in this sample population.

Cortisone in saliva of pigs: validation of a new assay and changes after thermal stress.

BACKGROUND: Cortisone is derived from cortisol through the action of the enzyme 11ß-hydroxysteroid dehydrogenase type II, and it has gained importance in recent years as a biomarker of stress. This study aimed to develop and validate an assay for the measurement of cortisone in pig saliva and evaluate whether its concentration varies in stressful situations. For this purpose, a specific immunoassay was developed and validated analytically, and a study was performed to evaluate whether cortisone concentrations in saliva can vary under heat stress conditions. RESULTS: The assay proved to be accurate, reliable, and sensitive for the measurement of cortisone in pig saliva. The limit of detection of the assay was set at 0.006 ng/ml, and the lower limit of quantification was 0.023 ng/ml. It also correlated significantly with the results obtained by LC‒MS/MS (P = 0.003; r = 0.64). In addition, the cortisone concentration in animals subjected to prolonged heat stress decreased significantly 15 days after treatment (P < 0.0001). CONCLUSIONS: According to these results, cortisone measured by this assay could be used as a tool for the non-invasive evaluation of thermal stress in pig saliva.

The use of serum paraoxonase-1 to assess inflammation in horses with colitis.

BACKGROUND: Paraoxonase-1 (PON-1) has been suggested as a marker of inflammation and oxidative stress in horses and could potentially be used for prognostication in horses with colitis. OBJECTIVES: Assessment of PON-1 in horses with colitis and comparison of two methods. METHODS: Serum PON-1 was measured by two methods (paraoxon and p-nitrophenyl acetate) in 161 horses with colitis and 57 controls. Follow-up samples obtained during hospitalization were available from 106 horses with colitis. The two methods were compared. RESULTS: Serum PON-1 was significantly lower in horses with colitis than in healthy horses (P < .0001 for both methods) as well as in nonsurvivors compared with survivors (P = .0141 [paraoxon-based method] and P < .0001 [p-nitrophenyl acetate-based method]), but with marked overlap between groups. PON-1 activity did not change parallel to a change in inflammatory status in response to treatment when assessed at admission and in up to seven follow-up samples. Admission PON-1 activity could not reliably classify horses as survivors or nonsurvivors, with sensitivity and specificity ranging between 53.1% and 72.9%. Results from the two methods were comparable. CONCLUSIONS: Both methods reliably measured serum PON-1 activity. Significant differences in PON-1 activity were found between healthy horses and horses with colitis and between survivors and nonsurvivors. However, PON-1 activity varied considerably within groups. Both the proposed reference intervals as well as alternative cutoff values resulted in suboptimal diagnostic and prognostic performance, and the use of serum PON-1 in horses with colitis thus seems to add little to existing diagnostic and prognostic markers.

Management of Long QT Syndrome: A Systematic Review.

Long QT syndrome (LQTS) is a cardiac disorder characterized by prolonged repolarization of the heart's electrical cycle, which can be observed as an extended QT interval on an electrocardiogram (ECG). The safe and effective management of LQTS often necessitates a multifaceted approach encompassing pharmacological treatment, lifestyle modifications, and, in high-risk cases, the implantation of implantable cardioverter-defibrillators (ICDs). Beta-blockers, particularly nadolol and propranolol, are foundational in treating LQTS, especially for high-risk patients, though ICDs are recommended for those with a history of cardiac arrest or recurrent arrhythmic episodes. Intermediate and low-risk patients are usually managed with medical therapy and regular monitoring. Lifestyle modifications, such as avoiding strenuous physical activities and certain medications, play a critical role. Additionally, psychological support is essential due to the anxiety and depression associated with LQTS. Left cardiac sympathetic denervation (LCSD) offers an alternative for those intolerant to beta-blockers or ICDs. For diagnosis and management, advancements in artificial intelligence (AI) are proving beneficial, enhancing early detection and risk stratification. Despite these developments, significant gaps in understanding the pathophysiology and optimal management strategies for LQTS remain. Future research should focus on refining risk stratification, developing new therapeutic approaches, and generating robust data to guide treatment decisions, ultimately aiming for a personalized medicine approach.

New insights into swine dysentery: faecal shedding, macro and microscopic lesions and biomarkers in early and acute stages of Brachyspira hyodysenteriae infection.

BACKGROUND: Swine dysentery (SD) is a severe mucohaemorrhagic colitis in pigs caused classically by Brachyspira hyodysenteriae. Although several aspects of B. hyodysenteriae infection dynamic are already described, further research in the early stage of this infection is required. In this study, 7-week-old pigs were orally challenged with B. hyodysenteriae to obtain information about faecal shedding, macro and microscopic intestinal lesions and serum acute phase proteins in pigs at the onset of B. hyodysenteriae shedding (early infection group, n = 8), in pigs with mucohaemorrhagic diarrhoea (acute infection group, n = 8) and in non-infected controls (n = 16). RESULTS: First B. hyodysenteriae detection by q-PCR and first loose stools with blood and mucus occurred both at 8 days post-inoculation. The lapse between a positive q-PCR and observation of mucohaemorrhagic diarrhoea ranged from 0 to 3 days, except in a single pig in which this period lasted 5 days. Macroscopic lesions were observed in the large intestine from both infected groups although more frequent and severe in acute infection group. Microscopic observation of the apex mucosa revealed that in early infection only higher ulceration values were observed compared to healthy controls. In contrast, the acute infection group exhibited higher ulceration, neutrophils infiltration and increased mucosal thickness compared to the other two groups. Among the serum biomarkers tested, only haptoglobin, C-reactive protein, and creatine kinase showed a significant increase in pigs in the acute infection period compared to controls, whereas haptoglobin was the only factor with a significant increase at the early infection compared to non-infected animals. CONCLUSIONS: This study provides new insights about SD and remarks the complex and limited options to perform an early detection of infected animals beyond PCR diagnosis.

Changes in saliva analytes in equine gastric ulcer syndrome (EGUS) after treatment: a pilot study.

Equine gastric ulcer syndrome (EGUS) is a prevalent condition in horses, affecting up to 93% of racehorses. Comprising the equine squamous gastric disease (ESGD) and the equine glandular gastric disease (EGGD), EGUS poses significant health challenges. Saliva, a non-invasive and easily obtainable sample, is increasingly recognized for its potential as a source of biomarkers in horses. This study investigates changes in saliva analytes using automated assays before and after EGUS treatment, aiming to identify biomarkers indicative of treatment success or failure. A total of 28 horses diagnosed with EGUS were treatment with omeprazole for six weeks and further divided into successful (n = 15) or unsuccessful (n = 13) treatment group. Saliva samples were collected before and after treatment, and analytes related to enzymes, metabolites, proteins, redox biomarkers, and minerals were measured using an automated chemistry analyzer. Results revealed that horses with successful treatment, indicated by reduced EGGD and ESGD scores, showed significant increases in bicarbonate and urea, and decreases in adenosine deaminase (ADA), and creatine kinase (CK). Conversely, horses with non-successful treatment showed no significant changes in salivary analytes. These analytes have the advantages of an easy and fast measurement and the possibility of being applied in routine. Further studies with larger populations should be performed to establish the possible practical application of these analytes as biomarkers of treatment.

Measurement of Trace Elements (Zinc, Copper, Magnesium, and Iron) in the Saliva of Horses: Validation Data and Changes in Equine Gastric Ulcer Syndrome (EGUS).

The objective of this study was to evaluate the possible use of spectrophotometric assays for the measurement of trace elements, including Zinc (Zn), Copper (Cu), Magnesium (Mg), and iron (Fe) in the saliva of horses and study their possible changes in equine gastric ulcer syndrome (EGUS). EGUS is a highly prevalent disease, with a current high incidence due to the increase in intensive management conditions. There are two EGUS diseases: equine squamous gastric disease (ESGD) and equine glandular gastric disease (EGGD), which can appear individually or together. For this purpose, automated spectrophotometric assays for measuring these analytes in horse saliva were analytically validated. Then, these analytes were measured in the saliva of horses with only ESGD, only EGGD, both ESGD and EGGD and a group of healthy horses. The methods used to measure the analytes were precise and accurate. Horses diagnosed with EGGD presented significantly lower levels of Zn and Mg. Fe concentrations were significantly lower in the saliva of horses with ESGD and EGGD. Overall, these results indicate that there are changes in trace elements in saliva in EGUS that could reflect the physiopathological mechanisms involved in this process and open the possibility of using trace elements as biomarkers of this syndrome.

Neutrophil-to-lymphocyte ratio and red blood cell distribution width to platelet ratio and their relationships with inflammatory and antioxidant status in dogs with different stages of heart failure due to myxomatous mitral valve disease.

We aimed to evaluate the red blood cell distribution width-to-platelet ratio (RDW/PLT) with other complete blood cell count (CBC) indices and their correlations with serum proinflammatory cytokines, acute phase proteins (APPs), and antioxidant biomarkers in dogs at different stages of heart failure (HF). A total of 29 dogs were divided into four groups according to the ACVIM Consensus Statement: stage-A (healthy/controls, n = 8), stage-B2 (n = 6), stage-C (n = 10), and stage-D (n = 5). Seventeen CBC indices were calculated and correlated with the measurements of inflammatory, APPs, and antioxidant biomarkers, as well as selected echocardiographic variables in all dogs. At stage-C, CBC indices were evaluated 14 days after the treatment. Statistically significant changes were observed only for RDW/PLT and neutrophil-to-lymphocyte ratio (NLR) between groups. NLR increased, but RDW/PLT deceased in dogs with HF, compared to controls (P < 0.05). There were no statistically differences between pre- and post-treatment CBC indices. There were significantly positive and negative correlations between the CBC indices, serum parameters and selected echocardiographic variables in dogs with HF(P < 0.05). ROC analysis showed the best sensitivity (57% and 68%) and specificity (100% and 57%) for NLR > 5.8 and RDW/PLT ≤ 0.057 for predicting the severity of HF, respectively. Results showed that NLR and RDW/PLT may have potential for monitoring severity of the disease and the effect of treatment in dogs with HF. Imbalances between indices of circulating blood cells can contribute to immunoinflammatory and antioxidant responses in pathogenesis of canine HF, which may provide us alternative targets to develop new diagnostic and therapeutic strategies in veterinary medicine.

Cystatin C, Ammonia, and Bicarbonate Measurements in the Saliva of Pigs: Analytical Validation and Changes in S. suis Infection.

Cystatin C, ammonia, and bicarbonate have been described to be biomarkers of sepsis and inflammation in humans. The saliva of pigs can be used to detect a wide range of pathogens but also many biomarkers that can be analyzed to evaluate different conditions such as stress (i.e., cortisol and alpha amylase), immune system (i.e., ADA, S100 proteins), inflammation (i.e., acute phase proteins), redox status (i.e., various antioxidants and oxidants), and general metabolism or the status of different organs and tissues. However, there is a lack of assays for the possible measurement and use of cystatin C, ammonia, and bicarbonate in saliva as biomarkers of sepsis or inflammation in pigs. The objective of this study was to validate commercially available automated assays for the measurement of cystatin C, ammonia, and bicarbonate in the saliva of pigs, having the advantage of using a noninvasive sample that is easy to collect. The assays were precise and accurate, and the recommended storage condition for the saliva samples was -80 °C. In addition, cystatin and ammonia showed significant increases in the saliva of pigs with S. suis infection, whereas bicarbonate decreased. Further studies would be recommended to increase knowledge about the possible potential applications of the measurements of these three analytes in the saliva of pigs as biomarkers to evaluate the animals' health and welfare.

Salivary biomarkers as pioneering indicators for diagnosis and severity stratification of pediatric long COVID.

Introduction: Long COVID, or post-acute sequelae of SARS-CoV-2 infection (PASC), manifests as persistent and often debilitating symptoms enduring well beyond the initial COVID-19 infection. This disease is especially worrying in children since it can seriously alter their development. Presently, a specific diagnostic test or definitive biomarker set for confirming long COVID is lacking, relying instead on the protracted presence of symptoms post-acute infection. Methods: We measured the levels of 13 biomarkers in 105 saliva samples (49 from children with long COVID and 56 controls), and the Pearson correlation coefficient was used to analyse the correlations between the levels of the different salivary biomarkers. Multivariate logistic regression analyses were performed to determine which of the 13 analysed salivary biomarkers were useful to discriminate between children with long COVID and controls, as well as between children with mild and severe long COVID symptoms. Results: Pediatric long COVID exhibited increased oxidant biomarkers and decreased antioxidant, immune response, and stress-related biomarkers. Correlation analyses unveiled distinct patterns between biomarkers in long COVID and controls. Notably, a multivariate logistic regression pinpointed TOS, ADA2, total proteins, and AOPP as pivotal variables, culminating in a remarkably accurate predictive model distinguishing long COVID from controls. Furthermore, total proteins and ADA1 were instrumental in discerning between mild and severe long COVID symptoms. Discussion: This research sheds light on the potential clinical utility of salivary biomarkers in diagnosing and categorizing the severity of pediatric long COVID. It also lays the groundwork for future investigations aimed at unravelling the prognostic value of these biomarkers in predicting the trajectory of long COVID in affected individuals.

The Role of the Insula in Chronic Pain and Associated Structural Changes: An Integrative Review.

Chronic pain affects a substantial portion of the global population, significantly impacting quality of life and well-being. This condition involves complex mechanisms, including dysfunction of the autonomic nervous system, which plays a crucial role in pain perception. The insula, a key brain region involved in pain processing, plays a critical role in pain perception and modulation. Lesions in the insula can result in pain asymbolia, where pain perception remains intact but emotional responses are inappropriate. The insula is anatomically and functionally divided into anterior and posterior regions, with the posterior insula processing nociceptive input based on intensity and location before relaying it to the anterior insula for emotional mediation. Understanding the insula's intricate role in pain processing is crucial, as it is involved in encoding prediction errors and mediating emotional dimensions of pain perception. The focus of this review was on synthesizing existing literature on the role of the insula in chronic pain and associated structural changes. The goal was to integrate findings from various sources to provide a comprehensive overview of the topic. The search strategy included a combination of Medical Subject Headings (MeSH) and relevant keywords related to insula and chronic pain. The following databases were surveyed: PubMed, Embase, Scopus, and Web of Science. We identified a total of 2515 articles, and after following the Preferred Reporting Items for Systematic Reviews and Meta-Analyses (PRISMA) guideline based on eligibility criteria, 46 articles were used to synthesize this review. Our study highlights the pivotal role of the insula in chronic pain processing and associated structural changes, integrating findings from diverse studies and neuroimaging investigations. Beyond mere pain sensation, the insula contributes to emotional awareness, attention, and salience detection within the pain network. Various chronic pain conditions reveal alterations in insular activity and connectivity, accompanied by changes in gray matter volume and neurochemical profiles. Interventions targeting the insula show promise in alleviating chronic pain symptoms. However, further research is needed to understand underlying mechanisms, which can aid in developing more effective therapeutic interventions for pain.

Overview of Modern Commercial Kits for Laboratory Diagnosis of African Swine Fever and Swine Influenza A Viruses.

Rapid and early detection of infectious diseases in pigs is important, especially for the implementation of control measures in suspected cases of African swine fever (ASF), as an effective and safe vaccine is not yet available in most of the affected countries. Additionally, analysis for swine influenza is of significance due to its high morbidity rate (up to 100%) despite a lower mortality rate compared to ASF. The wide distribution of swine influenza A virus (SwIAV) across various countries, the emergence of constantly new recombinant strains, and the danger of human infection underscore the need for rapid and accurate diagnosis. Several diagnostic approaches and commercial methods should be applied depending on the scenario, type of sample and the objective of the studies being implemented. At the early diagnosis of an outbreak, virus genome detection using a variety of PCR assays proves to be the most sensitive and specific technique. As the disease evolves, serology gains diagnostic value, as specific antibodies appear later in the course of the disease (after 7-10 days post-infection (DPI) for ASF and between 10-21 DPI for SwIAV). The ongoing development of commercial kits with enhanced sensitivity and specificity is evident. This review aims to analyse recent advances and current commercial kits utilised for the diagnosis of ASF and SwIAV.

Validation of two immunoassays for oxytocin measurements in human saliva.

The objective of this research was to develop and validate two immunoassays for oxytocin measurement in human saliva, one using a monoclonal and the other a polyclonal antibody against oxytocin, whose affinity for oxytocin was tested by an antibody mapping epitope analysis. These assays were analytically validated and used to compare oxytocin concentrations with those obtained with a commercial kit before and after the extraction or reduction/alkylation (R/A) treatments to saliva samples. The assays were also used to evaluate changes in salivary oxytocin concentrations following a physical effort and an induced psychological stress, which have previously been described as situations that cause an increase in salivary oxytocin. Both assays showed to be precise and accurate in the validation studies, and the antibodies used showed a defined binding region in case of the monoclonal antibody, whereas the polyclonal antibody showed binding events through all the oxytocin sequence. Although the monoclonal and polyclonal assays showed a positive correlation, they give results in a different range of magnitude. Both assays showed significant increases in oxytocin concentrations when applied after the physical effort and the psychological stress. This study shows that a variability in the reported values of oxytocin can occur depending on the assay and indicates that the use of different types of antibodies can give a different range of values when measuring oxytocin in saliva.

Comparison of the Effect of Two Different Handling Conditions at Slaughter in Saliva Analytes in Pigs.

In this report, different handling conditions at slaughterhouse were studied to assess changes in salivary biomarkers. For this purpose, finishing pigs were divided into two groups, one in which handling was improved to minimize stress (Group A, n = 24, transported and stabled at the slaughterhouse at low density without mixing with unfamiliar animals throughout the whole process) and another one in which animals had a more stressful handling process (Group B, n = 24, transported and stabled at high density with unfamiliar animals). Saliva samples were taken the day before transport to the slaughterhouse at 8:00 a.m. (B0) and 12:00 a.m. (B4), and the day of slaughter just after unloading animals at the slaughterhouse at approximately 8:00 a.m. (S0) and after 4 h of lairage at approximately 12:00 a.m. (S4). Group B showed significantly higher cortisol, total esterase activity, oxytocin, adenosine deaminase and haptoglobin levels than the Group A at both S0 and S4 sampling times, and higher levels of calprotectin and creatine kinase at S4 sampling time. This report indicates that differences in the way in which the pigs are handled at the slaughterhouse can lead to changes in salivary biomarkers and opens the possibility of the use of biomarker at slaughter to monitor handling conditions.

Evaluation of the presence of gingivitis as confounding factor in assessing inflammatory status in serum and saliva of dogs with diabetes mellitus.

The aim of this study was to evaluate the changes in the serum and salivary inflammatory markers induced by Diabetes mellitus (DM) in dogs and to assess the possible confounding effect of gingivitis. A panel of 13 cytokines was measured in the serum and saliva of dogs diagnosed with DM and compared with healthy dogs without gingivitis (control group 1; CG1) and dogs with gingivitis but otherwise healthy (control group 2; CG2). The results of the present study showed statistically significantly higher levels of IL-8, KC-like and MCP1 in the serum of dogs with DM compared to CG1 dogs. In the case of saliva, the DM group presented statistically higher GM-CSF, IL6, IL15, and MCP1 levels compared to CG1, and lower KC-like chemokine compared to CG2. Finally, gingivitis produced changes in saliva, with salivary levels of GM-CSF, IL-6, IL-7, IL-15, IP-10, KC-like, IL-10, IL-18, MCP1, TNFα being statistically significantly higher in the saliva of CG2 dogs compared to CG1. The results of the present study indicate that dogs with DM have altered cytokine levels in serum and saliva compared to healthy dogs. In addition, this study highlights the importance of taking oral health into account when determining cytokines in dogs, as gingivitis can significantly alter their concentrations. .

A systematic review and meta-analysis of the validation of serological methods for detecting anti-Toxoplasma gondii antibodies in humans and animals.

Toxoplasma gondii is a paradigmatic zoonotic parasite from the One Health perspective, since it is broadly distributed and virtually infects all warm-blooded species. A wide variety of serological techniques have been developed to detect T. gondii infection in humans and animals. Our aim was to describe and compare the main characteristics of these serological tests and validation processes and to critically analyze whether these tests meet the standards required to ensure an accurate serological diagnosis. The current systematic review and meta-analysis included 134 studies that were published from 2013 to 2023. QUADAS 2 tool was used to evaluate the quality of the included studies. A total of 52 variables related to the characteristics of the techniques and analytical and diagnostic validation parameters were studied. A wider panel of tests was developed for humans, including techniques exclusively developed for humans that involve costly equipment and the measurement of different Ig isotypes that are considered biomarkers of congenital toxoplasmosis. Studies conducted in humans frequently employed commercial techniques as reference tests, measured different immunoglobulin isotypes with a predominance for IgG (>50%) and discriminated between acute and chronic infections. In animals, the most commonly used reference techniques were in-house tests, which almost exclusively detected IgG. Common limitations identified in a large number of studies were some misunderstandings of the terms "gold standard" and "reference test" and the absence of information about the negative and positive control sera used or the exact cutoff employed, which were independent of the quality of the study. There is a lack of analytical validation, with few evaluations of cross-reactivity with other pathogens. Diagnostic odds ratio values showed that indirect ELISA based on native or chimeric antigens performed better than other tests. The reproducibility of serological test results in both humans and animals is not guaranteed due to a lack of relevant information and analytical validation. Thus, several key issues should be considered in the future, including interlaboratory ring trials.

Automated Spectrophotometric Assays for the Measurement of Ammonia and Bicarbonate in Saliva of Horses: Analytical Validation and Changes in Equine Gastric Ulcer Syndrome (EGUS).

Ammonia (NH3) and bicarbonate (HCO3) have been related to gastric ulcers in humans. Ammonia is considered a possible cause of gastric ulcers, whereas bicarbonate has a protective function. The presence of ulcers in the stomach of horses is defined as Equine Gastric Ulcer Syndrome (EGUS), which is a frequent disease in this species, and it has been associated with changes in saliva composition, such as in analytes related to inflammation, immune system and oxidative stress. The objectives of this study were (1) to perform an analytical validation of two automated spectrophotometric assays, one for ammonia and one for bicarbonate, in the horses' saliva and (2) to evaluate their possible variations with EGUS. Analytical validation of the automated assays for ammonia and bicarbonate in the saliva of horses showed that both assays were precise and accurate. In addition, significantly higher values of ammonia and lower values of bicarbonate were found in the saliva of horses with EGUS compared to healthy horses. It can be concluded that ammonia and bicarbonate can be measured in the saliva of horses and that ammonia increases and bicarbonate decreases in this sample type could be related to the presence of EGUS in this species.

Association between Eosinophil Count and Cortisol Concentrations in Equids Admitted in the Emergency Unit with Abdominal Pain.

Stress leukogram includes eosinopenia as one of its main markers (neutrophilia, eosinopenia, lymphopenia, and mild monocytosis). Cortisol is the main stress biomarker, which is also strongly correlated with the severity of gastrointestinal diseases. This study aimed to determine the relationship between salivary cortisol and the eosinophil cell count (EC) in equids with abdominal pain. To do this, 39 horses with abdominal pain referred to an emergency service were included. All samples were taken on admission, and several parameters and clinical data were included. Equids were classified according to the outcome as survivors and non-survivors. Non-surviving equids presented higher salivary cortisol concentrations (Non-Survivors: 1.580 ± 0.816 µg/dL; Survivors 0.988 ± 0.653 µg/dL; p < 0.05) and lower EC (Non-Survivors: 0.0000 × 103/µL (0.000/0.0075); Survivors: 0.0450 × 103/µL (0.010/0.1825); p < 0.01). In addition, the relationship between salivary cortisol concentration, EC, and the WBC was determined. Only a strong correlation (negative) was observed between cortisol and EC (r = -0.523, p < 0.01). Since cortisol is not an analyte that can be measured routinely in clinical settings such as emergencies, the EC could be a good alternative. While the results are promising, further studies are needed before EC can be used confidently in routine practice to predict survival in cases of abdominal pain.