PULLMAN, WA, JANUARY 16, 2023 —VMRD, Inc is excited to share the launch of their latest offering for equine veterinarians, a novel horse-side test to measure IgG in foals. VMRD’s Foal IgG test does not require refrigerated storage and runs in 10 minutes using the same palm-sized reader as the VMRD SAA test. The reader then objectively interprets the test and displays results, eliminating the stress of visual interpretation. “The Foal IgG test was designed with equine veterinarians in mind, incorporating features that address the challenges they face in the field,” according to Veterinary Affairs Manager Siddra Hines, DVM, PhD, DACVIM. “This allows vets to maintain their focus on the most important part of the job – their patients.” Failure of passive transfer in foals, characterized by low circulating IgG, can lead to devastating consequences by rendering the foal susceptible to a myriad of infections. Measuring IgG to ensure they absorbed enough antibody from the mare’s colostrum is therefore vital for any neonatal foal evaluation, helping the veterinarian determine whether treatment is indicated. “The Foal IgG product is a great example of VMRD’s vision of improving animal health through innovative and accurate diagnostics,” says CEO Ethan Adams. “This latest test in our equine point-of-care portfolio yields timely and accurate information pertaining to failure of passive transfer, improving foal health through better management and treatment decisions. That’s what we are in business to achieve!” For more information on VMRD point-of-care products, including VMRD Foal IgG and VMRD SAA, please visit our website at vmrd.com/poc or contact us at vets@vmrd.com. VMRD MEDIA CONTACTS: Siddra Hines, DVM, Ph.D, DACVIM Veterinary Affairs Manager 509-334-5815 siddrah@vmrd.com

It's every equine vet's favorite time of year - and I don't mean colic season! We're looking forward to seeing everyone next week at the Annual AAEP Convention in San Antonio and sharing our products that offer horse-side results for horse-side decisions. Come by booth # 7113 to learn about our user-friendly serum amyloid A (SAA) test with lot-specific calibration for maximum test performance, and take advantage of our AAEP promotion to save money on your reader! As a bonus, we are also excited to share our upcoming test for Foal IgG that requires no refrigeration and uses the same reader as VMRD SAA. No more stress trying to interpret color - the reader does the work for you! Keep scrolling for more show information & links. CONFERENCE SPECIAL Want to know what questions to ask us at our booth? Brush up on your VMRDSAA knowledge with our Equine SAA Educational Resources

Cats are notoriously stoic and hide illness well, often to their own detriment. This presents challenges for veterinarians when trying to diagnose and manage disease, as clinical signs are often subtle until the situation is critical. SAA is a biomarker to help diagnose infection in multiple species In recent years, serum amyloid A (SAA) has been extensively utilized by equine veterinarians with great success for the identification and monitoring of infection, showing greater sensitivity than fever for detecting subclinical infection.(1) SAA is the most sensitive acute phase protein in cats(2) as well as horses, therefore applicability of this valuable tool for feline medicine is quite similar. Measuring SAA facilitates early and effective detection of many infections, since it strongly mirrors clinical condition. SAA increases more predictably than WBC count(3,4) and may increase even in the absence of fever(1). It is also unaffected by NSAID or corticosteroid administration. SAA is normally almost zero(2), but increases rapidly and dramatically with acute, systemic inflammation.(3) As inflammation begins to resolve, it quickly returns to baseline level. Tracking SAA allows objective monitoring of treatment efficacy and can detect re-emergence of disease or complications before they become clinically evident. SAA increases with acute inflammation caused by bacterial or viral infection As an acute phase protein, SAA increases with bacterial or viral infections that cause acute, systemic inflammation. It is less affected by chronic or highly localized disease. A recent study at a primary care facility examined clinical use of SAA in cats for a variety of diseases(2), with results relevant for small animal practitioners. Feline SAA was most consistently and significantly elevated with upper respiratory infections, pneumonia, pyometra, feline infectious peritonitis, and traumatic disease. Other diseases in which SAA was more variable but still showed notable elevations included pancreatitis, hepatitis/cholangitis, ketoacidosis, and potentially gingivostomatitis. Patients with gastroenteritis, round cell tumors, lower urinary tract diseases, and hyperthyroidism occasionally had mild to moderate SAA elevation, but most were normal, as were the majority of patients with chronic kidney disease, solid tumors, cardiomyopathy, and diabetes mellitus (without ketoacidosis).(2) Monitoring clinical condition with SAA is particularly valuable A published case report demonstrating the clinical value of monitoring SAA tracked a cat with pancreatitis for over 2 years.(4) Upon initial presentation with acute pancreatitis, the cat had a 2-day history of anorexia and vomiting and was febrile with elevated SAA and normal WBC count. He improved markedly after 5 days of treatment and SAA returned to normal. In the ensuing follow-up, the cat experienced several acute exacerbations of chronic pancreatitis and other complications. In each instance SAA was again elevated, with variable (often normal) WBC results. SAA consistently returned to normal as clinical signs resolved.(4) Rapid, in-clinic SAA testing enhances patient management Coupled with other routine tests, measuring feline SAA in-clinic gives veterinarians an invaluable aid for differential diagnosis, early detection of infection, evaluating prognosis(5), and monitoring clinical condition. Tests can never replace a good physical exam or clinical expertise, but timely, objective results can make a difference for managing our reticent feline patients. REFERENCES 1 Oertly M, Gerber V, Anhold H, et al. The accuracy of serum amyloid A in determining early inflammation in horses after long-distance transportation by air. J Equine Vet Sci 2021;97:103337. DOI: 10.1016/j.jevs.2020.103337 2 Yuki M, Aoyama R, Nakagawa M, et al. A clinical investigation on serum amyloid A concentration in client-owned healthy and diseased cats in a primary care animal hospital. Vet Sci 2020;7(2):45-53. 3 Tamamoto T, Ohno K, Ohmi A, et al. Verification of measurement of the feline serum amyloid A (SAA) concentration by human SAA turbidimetric immunoassay and its clinical application. J Vet Med Sci 2008;70(11):1247-1252. 4 Tamamoto T, Ohno K, Ohmi A, et al. Time-course monitoring of serum amyloid A in a cat with pancreatitis. Vet Clin Pathol 2009;38(1):83-86. 5 Tamamoto T, Ohno K, Takahashi M, et al. Serum amyloid A as a prognostic marker in cats with various diseases. J Vet Diagn Invest 2013;25(3):428-432.

PULLMAN, WA, MARCH 01, 2022 – VMRD, a leading veterinary diagnostics manufacturer, is pleased to announce the release of their new test, VMRD SAA for Feline. “In recent years, SAA has been used extensively by equine veterinarians with great success to help diagnose infection and monitor treatment efficacy,” says Veterinary Affairs Manager Siddra Hines, DVM, PhD, DACVIM. “Following the release of VMRD SAA for Equine in 2019, we realized there was an unmet need for such a valuable tool in feline medicine as well.” SAA is an inflammatory biomarker that increases rapidly and dramatically with most infections, which can be particularly challenging to diagnose in cats. It is often more sensitive than fever or white blood cell count for detecting subclinical infections and decreases rapidly when effective treatment is instituted. “Cats are very stoic creatures, and often do not show signs of serious illness until the situation is critical,” states Chief Scientific Officer Don Knowles, DVM, PhD, DACVP. “Now VMRD SAA for Feline can help companion animal veterinarians identify infections early and objectively monitor clinical condition in cats.” VMRD SAA for Feline is a 10-minute test that can be run in-clinic on a handheld reader, which provides numerical results for easy interpretation. For more information on VMRD point-of-care products, including the VMRD SAA test for Feline, please visit our website or contact us at vets@vmrd.com.

Catching up with Dr. Boutros Bringing SAA on Board for Equine Practice In this continuing feature, we catch up with Dr. Christopher Boutros from Waller Equine Hospital in Waller, TX. Waller Equine Hospital is a full service primary care and referral hospital, offering a wide range of services in surgery, medicine, reproduction, emergency, preventative care, and general health. Dr. Boutros and his team have used Coggins tests for many years, and became early adopters of VMRD Equine SAA shortly after its release. What was your initial general impression of SAA testing, in general? "Quick and convenient. Best diagnostic tool for investigating or monitoring an infectious process." What is an example of one thing you were wrong or worried about? "There was some hesitation about incorporating a new test. However, it is easy to run and interpret and at this point it would be hard to practice without SAA testing." How have you incorporated in your day-to-day? "Neonatal and foal screening, differentiating allergic airway disease from respiratory infections, investigating fevers or ‘ADR’ (Ain't Doing Right) horses, investigating infections and monitoring antibiotic therapy efficacy and duration of treatment." Can you provide an example of how a normal result has been beneficial? "Helps to determine if a horse has heaves or a respiratory infection and helps guide length of antibiotic therapy." What is something that you appreciate about VMRD's offering? "Compact size, easy to operate, more accurate than other systems." What would you like others to know who might be on the fence on incorporating SAA testing to their practice? "Just do it! More helpful than a complete blood cell count. SAA increases dramatically in infectious cases, and it helps clients understand the importance of the treatment plan. It improves client compliance. What do you like most about VMRD's Equine SAA Test and/or Reader? "Very reliable results. Fast and simple to use. What do you like most about working with VMRD? "VMRD Has great quality tests and great people. VMRD offers great support for their products. They are a resource we can rely on." Anything else you would like to add? "We really appreciate the dedicated staff. VMRD is more than just a vendor, it is a company that we rely on and have a relationship with." Want to connect with Dr. Boutros and his team? You can find them at: Waller Equine Hospital 28234 FM 2920 Rd Waller, TX 77484 Phone: 936-931-2900 Website: wallerequine.com Facebook: https://www.facebook.com/Waller-Equine-Hospital-111150545581683/

Catching up with Dr. Torrisi Bringing SAA on Board for Equine Ambulatory Practice In this feature, we catch up with Dr. Kelly Torrisi, owner of Auburn Equine in Auburn, CA. Dr. Torrisi’s clinic employs 3 vets as well as 4 assistants and admins. On an average day, Auburn’s team attends to equine patients throughout the region, providing care from routine wellness checks on up to 24-hour emergency services. Dr. Torrisi and her team were one of VMRD’s early adopters of serum amyloid A testing and are now loyal users. We wanted to get an inside look at what that was like for her and her practice to bring a new, novel test on board. What was your initial general impression of SAA testing? "Initially, I didn’t see the need. I was doing fine without it." What concerns did you have about SAA that ended up not being a problem? "That it’s too expensive and my clients wouldn’t want to spend the money on the test and would rather spend the money on the treatments. Actually, clients don’t want to give medications if they don’t have to and I haven’t had many clients decline the SAA if offered." How have you incorporated SAA into your patient care? "I find that using the VMRD SAA test has been a huge compliment to my practice. The cost is not prohibitive for clients, and it has helped my case management to define a better treatment plan. There have been a few cases where the patient has not had a clear diagnosis, and there were financial constraints for the client, and I was able to make a good decision within 10 minutes with the client and patient present." How have normal results been beneficial? "It’s a handy rule in or rule out tool on the farm. Clients haven’t been mad when the results are normal, they are appreciative to know how to proceed with treatments." What benefit of VMRD's product do you feel appeals to smaller clinics in particular? "I appreciate the ability to not buy too many and risk the cartridges expiring, so far, I haven't had one expire!" What would you like others to know who may be considering incorporating SAA testing in their practice? "As someone who was one of the holdouts on SAA for many years and finally decided to give it a try, I feel as though it has improved my quality of medicine with prescribing medications in a timely manner as well as improving my assessment of treatment for ongoing medical cases." Want to connect with Dr. Torrisi and her team? You can find them at: Auburn Equine 1507 Grass Valley Hwy Auburn, CA 95603 Phone: 530-823-0162 Email: auburnequine@gmail.com Website: https://www.auburnequine.com/ Facebook: https://www.facebook.com/AuburnEquine/

Serum amyloid A (SAA) testing is a valuable tool for equine practitioners, whether they work in ambulatory practice or a referral hospital. Understanding the nature and dynamics of SAA allows testing to be utilized to its best potential, and proper case selection is important. Interpreting results in light of other clinical information is also vital. Collecting the right samples at the right time and using the correct technique for testing ensures that results will be useful and consistent. When measuring a single value, exact numerical results are less important as long as they are in the same clinical range (i.e. lead to the same clinical decision). However, consistency in technique becomes more important for effective monitoring. SAA is particularly valuable for differentiating infection SAA is most valuable in cases where infection causing acute, systemic inflammation is of concern. Although bacterial and viral infections both cause elevated SAA, it tends to be much more pronounced with bacterial. Importantly, SAA can help differentiate between acute infections (such as bacterial pneumonia) and other chronic or allergic etiologies (such as equine asthma) that may have similar clinical presentation. This facilitates appropriate and judicious administration of antimicrobials or other therapies. Results should of course be combined with a good physical exam, history, and other appropriate diagnostics to determine the nature of infection. It is also important to recognize that SAA will not increase significantly with chronic or localized issues. Potential confounding factors, such as recent vaccination, should be considered when evaluating results (see Unexpected Results article). Results from EDTA blood can remain consistent up to 7 days after collection Fresh blood should be tested immediately before it begins to clot. Anticoagulated blood samples can also be collected and tested later. We have demonstrated clinically consistent results with the VMRD SAA test up to 7 days after collection using EDTA blood samples. Blood samples can be stored refrigerated or at room temperature but should not be frozen. If blood has been stored, it should be resuspended thoroughly but gently prior to testing to break up rouleaux formation. The necessary dilution for serum/plasma differs from whole blood, therefore the appropriate testing protocol and sampling materials should be used. Although hematocrit does not appear to affect SAA results, there can be slight differences between fresh blood, anticoagulated blood, serum, and plasma due to other unknown factors. Diluted samples are stable for at least 30 minutes When collecting blood using the capillary, it is best to avoid bubbles in the capillary tube and avoid transferring extra blood on the outside of the tube. However, neither of these situations should change the results drastically. Once diluted, the sample should be stable in the diluent for at least 30 minutes. After 1-2 hours the value may decrease slightly but should still remain relatively accurate up to 24 hours for all but the lowest SAA concentrations. Vigorous horizontal shaking mixes samples thoroughly Tests should be run on a flat surface to ensure even flow of the sample through the test membrane. Particularly in high humidity environments, the test cartridge should be used within 10 minutes of opening, as exposure to moisture in the air can also affect flow. Samples should be mixed thoroughly by holding the assembled dropper horizontally and shaking vigorously. Although this may cause the formation of some bubbles, they should not interfere with the test. When dispensing the diluted sample, discard several drops prior to applying the solution to the sample well to ensure there is no plain diluent or bubbles in the dropper channel. Drops should be applied holding the dropper vertically. If less than 3 drops are applied the test may not run properly but adding more than 3 drops should not cause any significant issues. In essence, if drop count is in doubt, add another drop. Using the lot-specific calibration card enhances accuracy of results Using the correct calibration card for the test lot being used will ensure that results are as accurate and consistent as possible. That being said, if the calibration card lot number does not match the test lot number, results will likely be clinically accurate but not as precise. Results should be read at exactly 10 minutes as they may change gradually, usually increasing slightly over time. Consistency is important for patient monitoring Keeping sample type consistent when monitoring a patient over time avoids any variation that could occur due to slight sample type differences. If possible, it is also best to test in similar environmental conditions and within a similar time frame after sample collection for best consistency. Whenever possible, the same lot of tests should be used for monitoring an individual patient, although the effect of lot-to-lot variation is minimized by the lot-specific calibration card that standardizes test performance over time. For guidance on appropriate timing of sample collection for patient monitoring, see our article focused on this topic (Monitoring article). During antimicrobial treatment, SAA can be used to evaluate efficacy and help determine the necessary duration of therapy. SAA will continue to increase for 2-4 days after an acute inflammatory insult but should begin to decrease within a few days of instituting effective therapy.

Early detection of disease is critical in neonates Foals are highly vulnerable and delicate compared to adult horses, making timely identification of disease particularly important for early intervention. Serum amyloid A (SAA) is a very sensitive biomarker for inflammation, especially that caused by infection, and can be tested horse-side to help identify problems early. Elevated SAA in neonatal foals can serve as an early indicator of potentially critical issues such as sepsis, respiratory disease, or diarrheal disease.(1,2) Test SAA as a part of standard neonatal health exams Normal foals can have their SAA checked at the time they are tested for IgG. This allows early screening for any developing issues and provides a baseline for comparison if any clinical abnormalities do develop. If SAA is mildly elevated (20-100 µg/mL) this may be normal, as it can increase post-parturition/post-suckling.(1) However, they should be retested within 12-24 hours to ensure SAA is decreasing back to normal (<20 µg /mL). Abnormal foals should be tested immediately upon presentation, and treatment considered even if only mild SAA elevation is observed (20-100 µg /mL).(1,2) If SAA is >100 µg /mL (or increasing) in any foal, further diagnostics and treatment should be pursued as an underlying condition is likely.(1) SAA is more reliable than fibrinogen or WBC count as a predictor of sepsis Sepsis is one of the most problematic and urgent conditions in equine neonatal medicine. Rapid intervention is critical, therefore treatment is often based on presumptive diagnosis. Since SAA can be tested at point-of-care, it provides rapid, vital input when evaluating differential diagnoses and instituting appropriate therapy. It is reliably elevated in almost all foals with bacterial sepsis, unlike WBC/neutrophil count which may be low, high, or normal depending on the nature of sepsis.(2) Fibrinogen may also be more unpredictable as it decreases with consumption of clotting factors, which can occur in severe disease.(2) WBC count and fibrinogen are still important basic parameters, however their potential variability also makes them less than ideal for monitoring treatment effectiveness. SAA can be particularly valuable in this respect, as it will return to normal rapidly as foals recover. WBC count and fibrinogen generally take more time and are more unpredictable.(2) SAA improves R. equi screening protocols Newer research has shown added value of incorporating SAA into screening protocols on farms endemic for Rhodococcus equi. On these farms, ultrasound screening is often used to identify foals with subclinical disease for proactive antimicrobial therapy. This has greatly improved the ability of veterinarians to detect any potentially affected foals and prevent clinical disease. However, about 80% of foals with subclinical ultrasound lesions never progress to clinical R. equi(3), even if left untreated. Further refinement of screening strategies would help decrease unnecessary antimicrobial use and the development of further antimicrobial resistance. When used as part of such a screening program, SAA can help predict progression in foals with lesions on ultrasound.(3) Some earlier studies found that SAA alone did not reliably identify foals with subclinical disease, perhaps due to the chronic and localized nature of early R. equi abscesses.(4) However, when incorporated with ultrasound for on-farm screening, SAA improves the ability to identify foals that will progress to clinical illness versus those that should resolve without treatment, allowing more targeted use of antimicrobials. Hyperimmune plasma products may contain SAA Results of SAA testing following administration of hyperimmune plasma should be interpreted with caution. Hyperimmune plasma products may contain a significant level of SAA which can affect systemic levels in foals.(5) This is generally limited to mild elevations (<100 µg/mL) post-administration and would be expected to decrease rapidly if there is no driving inflammatory stimulus for production by the foal itself. Rechecking SAA 24 hours later should reveal an identifiable drop in value in normal foals, and of course results should always be considered in light of physical exam and other clinical findings. It is unknown whether normal commercial plasma contains any significant level of SAA, but it is less likely since donors are not standardly receiving repeated inoculations that would stimulate inflammation as with hyperimmune products. SAA testing can improve foal care with early, horse-side detection of inflammation caused by infection Testing SAA gives veterinarians a horse-side tool to detect issues sooner and treat our most vulnerable patients faster and more effectively. Ultimately, diagnosis and treatment decisions are made through clinical judgement, based on the available evidence. SAA provides valuable input for this process, leading to improved medical care and foal health. REFERENCES 1 Stoneham SJ, Palmer L, Cash R, et al. Measurement of serum amyloid A in the neonatal foal using a latex agglutination immunoturbidometric assay: determination of the normal range, variation with age and response to disease. Equine Vet J 2001;33(6):599-603. 2 Hultén C, Demmers S. Serum amyloid A (SAA) as an aid in the management of infectious disease in the foal: comparison with total leucocyte count, neutrophil count and fibrinogen. Equine Vet J 2002;34(7):693-698. 3 McCracken JL. Evaluation of white blood cell, fibrinogen, serum amyloid A, and ultrasonographic grade to refine an R. equi screening program in Proceedings. Am Assoc Equine Pract 2019;65:522-530. 4 Passamonti F, Vardi DM, Stefanetti V, et al. Rhodococcus equi pneumonia in foals: An assessment of the early diagnostic value of serum amyloid A and plasma fibrinogen concentrations in equine clinical practice. Vet J 2015;203(2):211-218. 5 Buchanan B. Effect of hyperimmune plasma administration on SAA blood values in foals. Texas Eq Vet Assoc "The Remuda" 2020;25:16-17. http://read.uberflip.com/i/1207332-2020-winter-the-remuda/13?, accessed 2/23/2021.

SAA should remain normal during pregnancy With normal conception and pregnancy, SAA level in broodmares is comparable to that in other healthy horses. That includes the time of ovulation through early pregnancy(1) and throughout the final trimester up to parturition(2,3). Artificial insemination may induce negligible SAA production, but levels stay within normal physiological limits. Although the majority of SAA is produced by the liver, local production by the endometrium likely contributes to systemic SAA elevation in pathological states.(5) Elevation early in pregnancy may indicate an increased risk of early embryonic death (EED), whereas any spike in mid-to-late-term should raise concern for ascending placentitis and abortion risk. In clinical application, other potential causes of increased SAA must of course also be considered, as elevation can occur due to other infections or pathological conditions.(2,6) However, SAA should remain normal in an otherwise healthy pregnant mare. Test SAA at ovulation and 7-14 days later to assess risk of EED Elevated SAA at the time of ovulation reflects a greater risk of experiencing EED, possibly due to some degree of subclinical endometritis. EED may occur in these individuals despite the continued presence of an intact corpus luteum and maintenance of normal progesterone levels.(1) In other mares with EED, SAA may be normal at ovulation but increase as early as 3 days post-ovulation. These cases are generally associated with excessive PGF2 and subsequent lysis of the corpus luteum, leading to EED. SAA typically remains elevated for at least 2 weeks, and sometimes up to 2 months.(1) For greatest utility in early pregnancy, SAA should be evaluated around the time of ovulation and then rechecked at 7-14 days. At either of these timepoints, even mild elevations of SAA (20-100 µg/mL) should be considered significant and warrant closer evaluation and monitoring. SAA can help identify mares at higher risk of abortion from ascending placentitis Monitoring SAA in mares with a history of high-risk pregnancy can provide valuable information, as it increases rapidly with ascending placentitis. This is in contrast to fibrinogen or white blood cell count, which do not appear to be useful markers for this condition.(3) Following a diagnosis of placentitis, tracking SAA can help monitor risk of impending abortion.(2,3) It increases on average 2-6 days prior to abortion from ascending placentitis and continues to rise until abortion occurs.(2,3) Treatment should lead to a decrease in SAA if the pregnancy is effectively preserved, and persistent elevation indicates a high likelihood of impending abortion.(2) SAA should return to normal within 2-3 days after foaling Normal parturition is an acute inflammatory event, and as such may increase SAA to a mild degree (<100 µg/mL). However, after normal foaling it should return to normal within 2-3 days.(2) More drastic or persistent elevation may indicate a medical issue requiring further investigation and care. REFERENCES 1 Krakowski L, Krawczyk CH, Kostro K, et al. Serum levels of acute phase proteins: SAA, Hp and progesterone (P4) in mares with early embryonic death. Reprod Dom Anim 2011;46:624-629. 2 Coutinho da Silva MA, Canisso IF, MacPherson ML, et al. Serum amyloid A concentration in healthy periparturient mares and mares with ascending placentitis. Equine Vet J 2013;45:619-624. 3 Canisso IF, Ball BA, Cray C, et al. Serum amyloid A and haptoglobin concentrations are increased in plasma of mares with ascending placentitis in the absence of changes in peripheral leukocyte counts or fibrinogen concentration. Am J Reprod Immunol 2014;72:376-385. 4 Wojtysiak K, Ryszka W, Stefaniak T, et al. Changes in the secretion of anti-inflammatory cytokines and acute phase pro­teins in the uterus after artificial insemination in the mare. Animals 2020;10(12):2438. 5 Christoffersen M, Baagoe CD, Jacobsen S, et al. Evaluation of the systemic acute phase response and endometrial gene expression of serum amyloid A and pro- and anti-inflammatory cytokines in mares with experimentally induced endometritis. Vet Immunol Immunopathol 2010;138(1-2):95-105. 6 Canisso IF, Loux SC, Lima FS. Biomarkers for placental disease in mares. Theriogenology 2020;150:302-307. 7 Ball BA. Embryonic loss in mares. Incidence, possible causes, and diagnostic considerations. Vet Clin North Am Equine Pract 1988;4(2):263-290 8 Giles RC, Donahue JM, Hong CB, et al. Causes of abortion, stillbirth, and perinatal death in horses: 3,527 cases (1986-1991). Am Vet Med Assoc. 1993;203(8):1170-1175

Colic can be one of the most rewarding and the most frustrating problems in equine medicine. Most cases are straightforward, however those that are not pose a constellation of challenges when making decisions for management. In many research studies and real-world situations, SAA testing has demonstrated value for multiple key aspects of colic management. SAA can help assess disease severity In concert with physical examination parameters and other diagnostics, SAA can provide valuable information in the assessment of a colic case. SAA concentration at initial presentation can help define disease severity and differentiate between possible etiologies, as well as providing a baseline for later comparison as the horse’s condition changes. SAA values can also offer input on prognosis, potential cost of care, risk of complications, and potentially need for surgery.(1-7) Elevated SAA may be an indication for referral While over 90% of colic cases are relatively simple and can be treated on-farm, the decision to refer is often critical for others. Testing SAA on-farm can help determine if referral is advisable, as elevated SAA is indicative of etiologies for which a higher level of care is likely required.(5-8) Colic cases that can be resolved medically and do not have a primary inflammatory cause tend to have low to normal SAA values.(1) If a horse is medically managed, either on-farm or at a referral hospital, re-evaluation of SAA can help track clinical condition and resolution of disease. The greatest elevations are seen with inflammatory causes of colic Since SAA increases with acute, systemic inflammation, elevations will be seen with some etiologies and not with others. The highest levels of SAA in colic cases are found with acute colitis, enteritis, peritonitis, or other inflammatory issues(5,8-10) all of which are likely to require advanced care. Higher values are also associated with more severe disease, such as small intestinal lesions and SIRS.(1-3) This also means that SAA tends to be higher in cases that will cost more to treat. SAA can help with surgical decisions and monitoring In the absence of a primary inflammatory etiology such as colitis, SAA is more likely to be high in colicky horses requiring surgery versus those that can be medically managed.(1) However, it is important to understand that peracute, severe issues such as colon torsion may initially have normal SAA as it takes at least 6-12 hours to start increasing. Following colic surgery, mild to moderate SAA elevation is expected as a direct result of surgery and primary disease, generally peaking at 72 hours post-op. However, values should begin to gradually decrease starting at 96 hours post-op.(1-3) Patients should therefore be checked at 72 hours to establish peak SAA, then ideally every 24-48 hours thereafter to track resolution of inflammation. Horses that develop post-operative complications generally remain at peak levels, and overall tend to have higher SAA in the first 1-5 days following surgery than those without complications.(2,3) At a minimum, all colic patients should be re-tested prior to discharge, as SAA should be decreasing or normal by this time.(2-4) If it is still elevated, the horse should be thoroughly evaluated for surgical site infections(4), catheter-associated thrombophlebitis, occult pneumonia, or other complications. Colic is one of the most common issues encountered by equine practitioners, leading to many sleepless nights and stressful situations for both veterinarians and horse owners. Although SAA is not the only factor to take into account when considering the cause and treatment of colic, it provides an objective and valuable tool to aid in the decision-making process, leading to improved patient care. REFERENCES 1 Westerman TL, Foster CM, Tornquist SJ, et al. Evaluation of serum amyloid A and haptoglobin concentrations as prognostic indicators for horses with colic. J Am Vet Med Assoc 2016;248(8):935-940. 2 Daniel AJ, Leise BS, Burgess BA, et al. Concentrations of serum amyloid A and plasma fibrinogen in horses undergoing emergency abdominal surgery. J Vet Emerg Crit Care 2016;26(3):344-351. 3 De Cozar M, Sherlock C, Knowles E, et al. Serum amyloid A and plasma fibrinogen concentrations in horses following emergency exploratory celiotomy. Equine Vet J 2020;52(1):59-66. 4 Aitken MR, Stefanovski D, Southwood LL. Serum amyloid A concentration in postoperative colic horses and its association with postoperative complications. Vet Surg 2018; 1–9. https://doi.org/10.1111/vsu.13133 5 Pihl TH, Scheepers E, Sanz M, et al. Influence of disease process and duration on acute phase proteins in serum and peritoneal fluid of horses with colic. J Vet Intern Med 2015;29:651-658. 6 Pihl TH, Scheepers E, Sanz M, et al. Acute-phase proteins as diagnostic markers in horses with colic. J Vet Emerg Crit Care 2016;26(5):664-674. 7 Vandenplas ML, Moore JN, Barton MH, et al. Concentrations of serum amyloid A and lipopolysaccharide binding protein in horses with colic. Am J Vet Res 2005;66(9):1509-1516. 8 Westerman TL, Tornquist SJ, Foster CM, et al. Evaluation of serum amyloid A and haptoglobin concentrations as prognostic indicators for horses with inflammatory disease examined at a tertiary care hospital. Am J Vet Res 2015;76(10):882-888. 9 Viner M, Mazan M, Bedenice D, et al. Comparison of serum amyloid A in horses with infectious and noninfectious respiratory diseases. J Equine Vet Sci 2017;49:11-13. 10 Hultén C, Sandgren B, Skiöldebrand E et al. The acute phase protein serum amyloid A (SAA) as an inflammatory marker in equine influenza infection. Acta Vet Scand 1999;40:323-333.

Routine equine health exams and biosecurity strategies utilize screening methods to detect underlying performance-limiting issues and potential infectious disease concerns. The fundamental basis of this is a good physical exam with rectal temperature screening, however subclinical issues may still exist that can become problematic. Measurement of serum amyloid A (SAA) is a highly effective tool for this purpose and can identify horses with subclinical disease that would not otherwise be detected. Any elevation of SAA should be considered abnormal The normal range of SAA in a healthy horse is generally considered to be less than 20 μg/mL.(1,2) It becomes elevated in acute systemic inflammation, particularly due to viral or bacterial infections, with bacteria stimulating the greatest SAA production.(3-5) SAA is highly sensitive and specific for the presence or absence of systemic inflammation, more sensitive than WBC count or fibrinogen.(1,3, 6-8) For the purpose of biosecurity or health screening, any elevation of SAA should be considered abnormal and trigger further investigation.(2,7) SAA may increase even if body temperature is normal SAA can increase even in the absence of fever(9-10), making it exceptionally useful for screening purposes and to monitor health status. Unlike fever, it will not be significantly affected by NSAID therapy.(7) In one study, SAA was 97.1% sensitive and 97.2% specific to differentiate clinically abnormal horses (i.e. those who developed infections) from those that were normal when tested 24 hours after air transportation.(9) The presence of fever had a sensitivity of just 2.9% at the same time point, indicating that SAA could identify brewing infections earlier and allow more rapid clinical intervention.(9) SAA testing can help prevent and manage infectious disease outbreaks There is clear value in assessing SAA alongside rectal temperature to identify subclinical disease(2,3,9) as part of an efficacious health screening strategy. In terms of biosecurity, disease outbreaks may be prevented if horses are effectively screened prior to co-mingling at events or when introducing new horses to a resident population. If an active infectious disease outbreak is occurring, SAA can help monitor at-risk or exposed horses for development of disease. Horses that have become infected will have elevated SAA and can be handled appropriately, with additional diagnostics as indicated.(3,4) SAA can also be used to monitor populations that may be at increased risk due to age, stress, exposure, population density, or other factors.(4,8) This could include young horses in intense training(4), hospital populations(6), or horses undergoing long-distance travel(9). SAA can identify subclinical problems in outwardly healthy horses As a general health screening tool, SAA can be very useful to assess the health of a horse prior to surgery and to monitor for complications afterwards.(3, 10-12) Testing prior to transport may identify subtle abnormalities that have the potential to develop into bigger problems with shipping. Testing horses after transport(3, 9) allows early recognition of shipping-related infections. SAA screening can help detect underlying issues during any routine health examination, including pre-purchase or insurance exams. Although normal (negative) results do not rule out all concerns, a positive result indicates an active problem that should be investigated. Normal horses should have little to no SAA As a general rule, a truly normal horse should have virtually no circulating SAA. SAA increases minimally or not at all with stress, exercise(13,14) or anesthesia4 alone. Barring any confounding factors (see April 2020 newsletter), elevated SAA in an outwardly healthy horse should always be a trigger to look deeper into possible causes. REFERENCES 1 Witkowska- Piłaszewicz OD, Żmigrodzka M, Winnicka A, et al. Serum amyloid A in equine health and disease. Equine Vet J 2019;51(3):293-298. 2 Radcliffe RM, Buchanan BR, Cook VL, et al. The clinical value of whole blood point-of-care biomarkers in large animal emergency and critical care medicine. J Vet Emerg Crit Care 2015;25(1):138-151. 3 Jacobsen S, Andersen PH. The acute phase protein serum amyloid A (SAA) as a marker of inflammation in horses. Equine Vet Educ 2007;19(1):38-46. 4 Pepys MB, Baltz ML, Tennet GA. Serum amyloid A protein (SAA) in horses: objective measurement of the acute phase response. Equine Vet J 1989;21(2):106-109. 5 Hultén C, Demmers S. Serum amyloid A (SAA) as an aid in the management of infectious disease in the foal: comparison with total leucocyte count, neutrophil count and fibrinogen. Equine Vet J 2002;34(7):693-698. 6 Belgrave RL, Dickey MM, Arheart KL. Assessment of serum amyloid A testing of horses and its clinical application in a specialized equine practice. J Am Vet Med Assoc 2013;243(1):113-119 7 Hooijberg EH, van der Hoven R, Tichy A, et al. Diagnostic and predictive capability of routine laboratory tests for the diagnosis and staging of equine inflammatory disease. J Vet Intern Med 2014;38:1587-1593. 8 Anhold H, Candon R, Chan Di-Sien, et al. A comparison of elevated blood parameter values in a population of thoroughbred racehorses. J Equine Vet Sci 2014;34:651-655. 9 Oertly M, Gerber V, Anhold H, et al. The accuracy of serum amyloid A in determining early inflammation in horses following long-distance transportation by air, in Proceedings. Am Assoc Equine Pract 2017;63:460-461. 10 Jacobsen S, Jensen JC, Frei S, et al. Use of serum amyloid A and other acute phase reactants to monitor the inflammatory response after castration in horses: a field study. Equine Vet J 2005;37(6):552-556. 11 De Cozar M, Sherlock C, Knowles E, et al. Serum amyloid A and plasma fibrinogen concentrations in horses following emergency exploratory celiotomy. Equine Vet J 2020;52(1):59-66. 12 Aitken MR, Stefanovski D, Southwood LL. Serum amyloid A concentration in postoperative colic horses and its association with postoperative complications. Vet Surg 2018; 1–9. https://doi.org/10.1111/vsu.13133 13 Kristensen L. Buhl R, Nostell K, et al. Acute exercise does not induce an acute phase response (APR) in Standardbred trotters. Can J Vet Res 2014;78(2):97-102. 14 Cywinska A, Witkowski L, Szarska E, et al. Serum amyloid A (SAA) concentration after training sessions in Arabian race and endurance horses. BMC Vet Res 2013;9:91-97.

Early and accurate detection of septic arthritis/synovitis is critical to minimize damage to cartilage and other structures that can ultimately affect the horse’s return to function and long-term prognosis. By the time diagnosis is clear cut, a significant amount of damage may be present. Synoviocentesis carries risks, especially when performed in the field, and can complicate standard joint fluid analysis when repeated sampling is required. Additional tools to enhance identification of sepsis and differentiation from non-septic processes are of high value to facilitate appropriate intervention and monitor treatment response. Blood SAA can be used to identify and monitor synovial sepsis In healthy horses with normal joints, serum amyloid A (SAA) is usually undetectable in blood and synovial fluid. With septic pathology however, SAA will increase in both. Interestingly, this increase is often more rapid and dramatic in blood.(1)  Blood SAA is also positively correlated with synovial total protein and total nucleated cell count.(2)  This allows blood SAA to be used as a surrogate for synovial fluid in many cases, which is particularly advantageous when synoviocentesis is not possible or advisable. Collection of blood is comparatively easy, making it easy to use SAA for monitoring treatment efficacy and resolution of infection, while avoiding the complications of repeated synoviocentesis.(3) SAA is not elevated with non-septic conditions In non-septic synovitis SAA is generally undetectable in both blood and synovial fluid, even when other synovial fluid abnormalities are observed.(1,2,4) Unlike total protein concentration, synovial SAA does not increase significantly with repeated arthrocentesis, through-and-through joint lavage, or with arthroscopy alone.(5,6)  It also will not be significantly elevated with low or chronic inflammatory states such as osteoarthritis.(7) SAA in synovial fluid comes from both the liver and the synovial membrane The majority of SAA is produced in the liver, however there is some local production in the synovial membrane.(7)  The SAA in synovial fluid results from a combination of both these sources. If blood SAA is elevated due to another systemic inflammatory condition, some systemically produced SAA may filter into the synovial fluid.(4)  Infection of other neighboring structures (such as septic osteitis or other septic synovial structures) can also cause a mild increase in synovial SAA.(4)  However, barring the presence of other significant disease processes, elevated synovial SAA also supports a diagnosis of septic synovitis. As with all adjunct diagnostic tests, SAA results must be interpreted in light of other clinical factors and diagnostic results such as joint fluid analysis. Ideally, SAA quantitation should be performed prior to the institution of therapy when possible.(4)  If less than 24 hours have passed since the initial injury, SAA in blood and synovial fluid may still be low(3), although some studies have shown an increase in as little as 6-8 hours.(4,7)  If SAA concentration is normal and sepsis is still suspected, blood or synovial SAA should be retested 12-18 hours later. Importantly, serial monitoring of blood SAA can be particularly valuable to track clinical progress without subjecting the horse to repeated synoviocentesis.(3) The VMRD SAA test is fully validated for use with blood, serum, or plasma. Studies with synovial fluid are ongoing – please contact us if you are interested in participating. References 1 Ludwig EK, Wiese RB, Graham MR, et al. Serum and synovial fluid serum amyloid A response in equine models of synovitis and septic arthritis. Vet Surg 2016;45(7):859-867. 2 Robinson CS, Singer ER, Piviani M, et al. Are serum amyloid A or D-lactate useful to diagnose synovial contamination or sepsis in horses? Vet Rec 2017;181(16):425-429. 3 Haltmayer E, Schendenwein I, Licka TF. Course of serum amyloid A (SAA) plasma concentrations in horses undergoing surgery for injuries penetrating synovial structures, an observational clinical study. BMC Vet Res 2017;13:137-147. 4 Stack JD, Cousty M, Steele E, et al. Comparison of serum amyloid A measurements in equine synovial fluid with routine diagnostic methods to detect synovial infection in a clinical environment. Front Vet Sci 2019;6:325. 5 Sanchez-Teran AF, Bracamonte JL, Hendrick S, et al. Effect of arthroscopic lavage on systemic and synovial fluid serum amyloid A in healthy horses. Vet Surg 2016;45:223-230. 6 Sanchez-Teran AF, Bracamonte JL, Hendrick S, et al. Effect of repeated through-and-through joint lavage on serum amyloid A in synovial from healthy horses. Vet J 2016;210:30-33. 7 Jacobsen S, Niewold TA, Halling-Thomsen M, et al. Serum amyloid A isoforms in serum and synovial fluid in horses with lipopolysaccharide-induced arthritis. Vet Immunol Immunopathol 2006;110:325-330.