Haematopoietic Flashcards
In vitro flow rates through five different catheters intended for intravenous use in horses at two different heights
-Larger diameter catheters achieved faster fluid flow rates, with the 10-gauge catheter at 150 mm length showing the highest rates.
-Increasing the height of the fluid bag significantly increased flow rates, with an average rise of 25.4–28.6% when the bag was elevated from 150 cm to 200 cm.
-There was excellent consistency in flow rates across identical catheters of the same batch, with intraclass correlation coefficients ranging from 0.994 to 0.998.
A risk assessment of equine piroplasmosis entry, exposure and consequences in the UK
Although the risk of endemic EP establishment is low, the potential consequences are severe, affecting animal health, welfare, and the equine industry.
Risk of Entry:
-Subclinical carriers present a significant risk as they may enter undetected due to a lack of mandatory screening. (med)
-Entry risks from infected ticks are low due to limited UK tick populations capable of EP transmission. However, Dermacentor reticulatus, an EP-capable vector, is established in some UK regions. (low)
Four primary exposure routes are identified:
-Vector-borne transmission: Limited by the restricted distribution of competent tick species in the UK. Low
-Mechanical transmission: Contamination of needles and equipment can pose a high infection risk. Med
-Haematological transmission: The use of infected blood in transfusions carries a very high infection risk. Low
-Transplacental transmission: This is the most probable exposure route, with a high risk of neonates being born infected. Very High
Consequences of EP Establishment:
-Acute disease in equines can lead to high mortality rates, particularly in untreated animals.
-Chronic carriers serve as a persistent reservoir for transmission, complicating disease control.
-Economic impacts include productivity losses, increased management costs (e.g., tick control, surveillance), and restrictions on international equine trade.
The study emphasizes the need for improved screening and biosecurity measures:
-EP should be classified as a notifiable disease in the UK.
-Mandatory serological testing of equines before importation.
-Pre-import ectoparasite treatment and certification.
-Adopting these measures would align the UK with international standards set by the OIE.
Modelling the probability and impact of false-positive serology for Borrelia burgdorferi sensu lato
-The apparent seroprevalence of Borrelia burgdorferi sensu lato in horses without clinical signs of Lyme borreliosis in southern Belgium was 22%.
-The true seroprevalence was estimated at 11%, indicating that a significant proportion of positive ELISA results were false positives.
-Two-thirds of ELISA-positive results were likely false positives in this population.
-The specificity of the ELISA test was lower than expected when applied to asymptomatic horses, demonstrating potential limitations in clinical contexts.
-The poor performance of ELISA and similar screening tests (e.g., IFA) underscores the need for confirmatory testing (e.g., Western Blot) for accurate diagnosis.
-Cross-reactivity with other pathogens and differences in test performance across geographical regions were identified as contributing factors to diagnostic inaccuracy.
Implications for Antimicrobial Use:
-The reliance on positive ELISA results for diagnosing Lyme borreliosis has led to unnecessary antimicrobial treatments. Approximately 5% of antimicrobial use in equine veterinary practice in southern Belgium could be attributed to overtreatment following false-positive serology.
- Unnecessary antimicrobial use raises concerns about antimicrobial resistance and the risk of adverse events, including antimicrobial-associated diarrhea.
Veterinary Practice Trends:
-A survey revealed that 60% of veterinarians would prescribe antimicrobials based on a positive Borrelia serology result, even in the absence of specific clinical signs.
-This highlights a need for improved education among veterinarians regarding the limitations of serological tests and the interpretation of results.
Sero-molecular survey and risk factors of equine piroplasmosis in horses in Spain
- 740 asymptomatic horses
-cELISA 42.9% CFT 41%(Se 80%, Sp 96%),
-30.3% were EP positive by PCR. T equi > B caballi.
-Lower sensitivity of CFT, particularly for B. caballi (T. equi: 80.3%, B. caballi: 45.7%). CFT primarily detects IgM antibodies, indicative of early infections, whereas cELISA is more effective for chronic cases due to its focus on IgG antibodies. - Exposure to T. equi was significantly higher than to B. caballi and the highest (sero)prevalence was detected in the northern communities.
- Assocaite with sero+T. equi : Increasing horse age, presence of ticks and contact with cows
- Associated with B. caballi - tetanus vaccination, ticks and fairs attendance (no) were associated
–T. equi persists lifelong, while B. caballi persists up to four years
-Higher seroprevalence in northern and western regions (e.g., Extremadura), likely due to favorable tick habitats (warm, humid climates and rural settings).
Immune-mediated haemolytic anaemia and thrombocytopenia in 25 adult equids: 1997-2016
-Concurrent IMHA and more common than IMHA or IMTP alone.
-Horses with IMHA or IMTP had higher odds of being diagnosed with neoplasia compared to control horses (Odds Ratio = 4.5).
Hypotheses for this association include:
-Shared immune system defects, such as imbalances in T-cell activation.
-Clonal T-cell expansion causing direct cytotoxic effects and promoting autoantibody production.
-Overexpression of oncogenes linked to concurrent immune-mediated diseases and neoplasia.
-Lymphoma, especially T-cell lymphoma, was the most commonly diagnosed neoplasm among affected horses.
Prognostic Indicators:
-Horses with secondary immune-mediated diseases, particularly those associated with neoplasia
-BUN -igher BUN levels correlating with increased mortality.
-Platelet count was significantly lower in nonsurvivors
Disease Characteristics:
-The prognosis for primary immune-mediated diseases was relatively good, warranting aggressive treatment.
-Secondary immune-mediated diseases, especially those linked to neoplasia, had a poor prognosis.
-Concurrent IMHA and thrombocytopenia were associated with higher mortality rates compared to isolated IMHA or IMTP cases.
2 horses IMHA only, 8 IMTP only, 15 both. 9 primary, 16 secondary
-survival to discharge 60% Long term (6 months) 4/15
-Secondary conditions included neoplasia, bacterial infection, enteritis, drug, hepatopathy, autologous BM aspirate injection.
-Treatment: corticosteroids. up to 113 days. blood transfusion, azathioprine.
- neoplasaias: T cell lymphoma, large B cell lymphoma.
Effect of leukoreduction on the metabolism of equine packed red blood cells during refrigerated storage
-LR significantly impacts the metabolic phenotype of equine packed RBCs (pRBCs), with more substantial effects than storage duration.
-The process preserves ATP pools, which are crucial for maintaining RBC function, including membrane deformability and preventing hemolysis.
-LR mitigates oxidative stress in RBCs by reducing reactive oxygen species and protecting antioxidant mechanisms.
Storage Lesions in RBCs:
-Storage lesions refer to metabolic and structural alterations in RBCs that occur over time during refrigeration.
Non-leukoreduced (nLR) RBCs accumulate more metabolic byproducts such as carboxylic acids (e.g., succinate, fumarate, malate) and oxidative damage markers, exacerbating storage lesions.
-LR slows the degradation of critical molecules like ATP and reduces the buildup of glycolytic byproducts and purine metabolites, improving the metabolic stability of RBCs during storage.
-nLR RBCs exhibit increased glycolytic flux, leading to faster depletion of energy reserves (e.g., ATP) and higher oxidative stress.
-LR stabilizes redox balance by maintaining reduced glutathione levels and limiting the activation of oxidative pathways like the pentose phosphate pathway.
Lipid and Amino Acid Metabolism:
-LR prevents the accumulation of lipid peroxidation products, including long-chain fatty acids and their oxidative derivatives, which can induce inflammatory responses in transfusion recipients.
-Amino acid catabolism, particularly the breakdown of purines and the accumulation of polyamines (e.g., spermidine, spermine), is reduced in LR RBCs, highlighting its role in modulating immunological reactions.
Clinical Relevance:
-LR is already standard practice in human transfusion medicine due to its proven benefits, but its application in equine transfusion remains inconsistent.
-This study demonstrates that LR improves the quality of stored equine pRBCs, potentially reducing adverse transfusion reactions and enhancing transfusion efficacy and safety in horses.
Humoral antibody response of 10 horses after vaccination against African horse sickness with an inactivated vaccine containing all 9 serotypes in one injection
-The inactivated African horse sickness virus (AHSV) vaccine containing all nine serotypes induced high levels of ELISA and virus-neutralizing antibodies in horses following a three-dose immunization schedule (primary, booster after 4 weeks, and booster after 6 months). -Despite the efficacy, ELISA antibodies declined significantly 3-5 months post-booster, necessitating periodic re-vaccination to maintain immunity.
Comparison to Existing Vaccines:
-While live attenuated vaccines are widely used, they carry risks of reversion to virulence and reassortment with field strains. The inactivated vaccine eliminates these risks, making it safer.
-Historical use of inactivated vaccines during outbreaks in Spain, Portugal, and Morocco demonstrated effectiveness but required frequent administration due to shorter duration of immunity.
-The requirement for multiple doses increases the cost and logistical challenges of vaccination programs.
-The large volume of the vaccine (8 mL/animal) used in this study was highlighted as a drawback, with plans to reduce the dose in future research.
-Both intramuscular (IM) and subcutaneous (SC) routes were evaluated, with IM administration being recommended due to fewer local reactions. SC administration resulted in transient swelling at the injection site in all vaccinated horses, particularly after the annual booster.
Economic assessment of African horse sickness vaccine impact
-The economic impact of AHS was estimated to be US$95 million per annum, and this was mainly in endemic regions with domestic equine industries and involved in international trade.
-Investment required to bring a new AHS vaccine to market was estimated to be up to US$3.5 million, which was very small relative to the benefits estimated in this study.
-AHS outbreaks in non-endemic countries, such as the 2020 outbreak in Thailand, underscore the risk and potential widespread economic damage.
-Non-endemic countries face the dual risk of outbreaks and the financial burden of maintaining measures to prevent incursions.
Investment in a safer and more effective AHS vaccine is economically viable, particularly for:
=The South African equine industry involved in domestic and international activities.
=European equine industries at risk of AHS incursion. A single outbreak in Europe, or a reduction in transportation costs for horses from South Africa to Europe, would alone justify the vaccine investment.
Impact on Working Equids:
-The cost-benefit analysis (CBA) for working equids in endemic areas showed insufficient returns to justify investment solely based on their use.
-However, the analysis likely underestimates the economic and social contributions of these animals, which are critical to livelihoods in many regions.
Broader Benefits and Equity Concerns:
-Development of a new vaccine could enhance the resilience of communities reliant on working equids, though this benefit is hard to quantify.
-Equity of vaccine distribution is vital to ensure accessibility for both high-value horse sectors and communities dependent on working equids.
-Better protection in endemic areas could indirectly benefit non-endemic regions by reducing risks of disease spread.
Immunoreactivity of canine, feline, and equine D dimer with antibodies to human D dimer
There are differences in the pathophysiology of coagulopathies in horses which tend to develop thromboembolic, rather than haemorrhagic, complications of DIC, reflecting widespread thrombin activation accompanied by release of D-dimer from fibrin thrombi. Thus horses might have a lower threshold for production of D-dimer, and higher circulating levels in health
Horses were immunopositive in D2D polyclonal Ig, DD5 (Fibrinogen) but not DD44 monoclonal.
D2D is poorly sensitive in humans but appears to be sensitive in horses.
Multiple other human assays have demonstrated good clinical performance in dogs and horses- these are incompletely validated.
Transfer of naturally acquired specific passive immunity against Anaplasma phagocytophilum in foals in Southeastern Pennsylvania and Northern Maryland
Transfer of specific passive immunity to A. phagocytophilum occurred in 80% of foals born to seropositive mares and declined by 3 months of age
Three out of 20 foals seroconverted between 3 and 6 months of age. These had mild clinical signs.
High seropositivity (91%) for Anaplasma phagocytophilum was observed in clinically healthy pregnant mares in Southeastern Pennsylvania and Northern Maryland.
The prevalence of infection in ticks within this region is relatively low (1.7%–3.3%), suggesting that seropositivity does not always indicate active infection but might reflect past exposure.
Despite the transfer of maternal antibodies, A. phagocytophilum infection should still be considered in foals presenting with clinical signs of equine granulocytic anaplasmosis (EGA), especially after the waning of passive immunity.
Between 3 and 6 months of age, 15% of foals seroconverted, with some displaying clinical signs like fever and intestinal hypermotility, suggesting natural exposure to A. phagocytophilum.
No significant difference was observed in maternal antibody titers between pre-foaling and foaling timepoints, suggesting no substantial depletion of maternal IgG due to colostral transfer.
This finding contrasts with studies in other species where periparturient drops in IgG levels are noted.
Did not sample pre colostrum
Equine leptospirosis: Experimental challenge of Leptospira interrogans serovar Bratislava fails to establish infection in naive horses
-The study challenges the long-held belief that Leptospira interrogans serovar Bratislava is host-adapted and pathogenic in horses.
-NB: One strain administered by topical ocular and intraperitoneal injection- need to look at other strains of the serovar.
-All challenged horses developed agglutination antibodies against serovar Bratislava within three days post-inoculation. Significant cross-reactivity occurred with antibodies for other serovars, complicating the identification of the infecting strain. The findings support that cross-reactivity is likely due to shared genomic features among serovars, as confirmed by pangenomic analysis.
-None of the challenged horses developed persistent infection or significant clinical symptoms, even though mild immune responses (e.g., higher WBC, lymphocyte counts) were observed. PCR and culture consistently failed to detect leptospires in blood, urine, or tissues, suggesting the bacteria were rapidly cleared. Changes in biochemical parameters (e.g., elevated bilirubin, decreased iron) were within reference intervals and attributed to transient inflammation or other non-specific responses.
-The MAT test, though considered the “gold standard,” lacks specificity for identifying infecting serovars due to significant cross-reactivity. Misdiagnosis using MAT results could lead to inappropriate treatments, including unnecessary antimicrobial use or incorrect public health interventions.
-Earlier studies suggested a host-adapted nature of serovar Bratislava based on serological prevalence and association with clinical cases.This study, however, found no genetic confirmation of colonization or pathogenesis in horses, reinforcing the hypothesis that cross-reactivity drove prior assumptions.
Seroprevalence and evaluation of risk factors associated with seropositivity for Borrelia burgdorferi in Ontario horses
-The seroprevalence of B. burgdorferi on at least one test was 17% (91/551), though only 15 (16%) horses tested positive with both tests.
-A spatial cluster of cases was detected in Eastern Ontario.
-The odds of being seropositive for B. burgdorferi on the C6 ELISA were significantly increased when oak trees were present by pastures (OR = 7.3 (1.8-29.2), P = .005), while the odds were significantly decreased when
regular tick checks were performed (OR = 0.1 (0.01-0.7), P = 0.02).
- Test has 33-66% FP rate
-Agreement between the two diagnostic methods (C6 ELISA and Multiplex ELISA) was low (Kappa = 0.27), complicating definitive diagnosis of exposure.
Evaluation of new leptospiral antigens for the diagnosis of equine leptospirosis: An approach using pan-genomic analysis, reverse vaccinology and antigenic selection
Reverse vaccinology is computationally driven approach to vaccine development that starts with analyzing the genome of a pathogen to identify potential vaccine candidates. Unlike traditional vaccinology, which relies on culturing pathogens and isolating antigens, reverse vaccinology leverages bioinformatics tools to predict antigens that could stimulate an immune response.
Key Steps in Reverse Vaccinology
-Genome Sequencing:
-Antigen Prediction: Computational tools analyze the genome to identify proteins likely to be good vaccine candidates.
–Key criteria for selection include: Antigenicity, Localization, Essentiality, Non-Homology: Proteins unique to the pathogen to avoid cross-reactivity with host proteins.
-Selection of Targets: Proteins with specific properties (e.g., B-cell epitopes, molecular weight limits) are prioritized. Antigenic scoring tools like VaxiJen and epitope prediction algorithms (e.g., ABCPred) are often used.
-Cloning and Expression: The genes encoding the selected proteins are cloned into expression systems (e.g., E. coli) to produce recombinant proteins.
-Experimental Validation: The recombinant proteins are tested in vitro and in vivo for their ability to: Elicit an immune response. Provide protection in animal models. ELISA or other immunoassays are used to confirm the antigen’s reactivity with sera from infected or vaccinated subjects.
-Final Vaccine Development: Promising antigens are formulated into a vaccine, followed by preclinical and clinical trials to confirm efficacy and safety.
The microscopic agglutination test (MAT) is the current gold standard for leptospirosis diagnosis but suffers from:
-Low sensitivity (40%-60%).
-High specificity (95%-100%).
-Cross-reactivity among serovars, leading to difficulties in identifying specific infecting serovars.
-High costs, labor intensiveness, and the risk of infection to laboratory workers due to handling live organisms.
ELISA as a Promising Alternative: simplicity, speed, and stability of coated microwell plates.
Eight novel leptospiral antigens were identified and expressed successfully using reverse vaccinology and antigenic selection criteria.
The LA_0711 antigen demonstrated the highest sensitivity (99%) and specificity (88%), making it a strong candidate for improved diagnostic tests.
A multi-antigen ELISA approach using combinations of the most accurate antigens improved sensitivity and specificity.
Parallel testing combinations allowed for 100% sensitivity, although specificity was slightly reduced.
Ultrasonographic technique and appearance of presumptively normal clinically relevant lymph nodes in horses
Medial Retropharyngeal Lymph Nodes:
Transducer Position: Microconvex transducer placed transverse to the common carotid artery at the cranial third of the neck.
Landmarks: Located near the angle of the mandible, between the external and internal carotid arteries, superficial to the guttural pouch, and deep to the salivary glands.
Depth: 2–5 cm from the skin surface.
Cranial Deep Cervical Lymph Nodes:
Transducer Position: Linear transducer placed transverse on the jugular groove and moved caudally to visualize the occipital and maxillary vein convergence.
Landmarks: Found adjacent to the maxillary vein, below the parotid gland, and near the external jugular vein.
Depth: 1–4 cm from the skin surface.
Caudal Deep Cervical Lymph Nodes:
Transducer Position: Convex or linear transducer in a sagittal or parasagittal plane along the ventral trachea at the neck’s caudal third.
Landmarks: Positioned near the thoracic inlet, around the bifurcation of the common carotid arteries and external jugular veins.
Depth: 5–10 cm depending on the horse’s size and applied pressure.
Caecal Lymph Nodes:
Transducer Position: Convex transducer applied to the right paralumbar fossa.
Landmarks: Found along the lateral caecal band, near the lateral caecal artery and vein, parallel to the costal arch.
Depth: 5–10 cm from the skin surface.
Lymph Nodes of the Iliosacral Lymphocentre:
Transducer Position: Small curvilinear transducer placed transrectally to localize the aorta and its bifurcation at the sacral promontory.
Landmarks: Located along the internal iliac arteries, within 3 cm of the transducer.
Challenges: Visualization is affected by anatomical variability and operator experience.
Validation of an indirect in-house ELISA using synthetic peptides to detect antibodies anti-gp90 and gp45 of the equine infectious anaemia virus
-ELISAgp90/45 was performed using 243 EIA positive and 878 negative equid sera, and showed a diagnostic sensitivity of 99.59% [CI 97.73%-99.99%] and a diagnostic specificity of 90.32% [CI 88.17%-92.19%],
-Analytical sensitivity of the ELISAgp90/45 was 800 times greater than that of AGID test for positive sera and 400 times greater for weak positive sera.
-ELISAgp90/45 also showed optimal analytical specificity, since no cross-reactivity was detected with antibodies against other equine viruses.
-ELISAgp90/45 detected antibodies earlier (within 10-15 days post-infection) compared to the AGID test, which typically detects antibodies after 14 days and sometimes up to 180 days.
-The ELISA identified approximately 25% more EIAV-infected horses than the AGID test,
