Neonatal isoerythrolysis involving the Qc and Db antigens in a foal

Jennifer M. MacLeay, DVM, PhD, DACVIM

RBC. The hospital reported that this process would take several hours; therefore, the foal was given 1 L of whole blood from a nonrelated gelding donor housed on the farm while the mares RBC were being washed. The filly responded well to this first transfusion, and although the foal was still exercise intolerant, rectal temperature, heart rate, and respiratory rate returned to normal. Twenty-four hours after parturition, the foal received a second transfusion, which consisted of 500 ml of the mares washed RBC (PCV, 80%) resuspended in 500 ml of isotonic saline (0.9% NaCl) solution (final PCV, 40%). The foals PCV was 18% after this transfusion. At this time, results of physical examination were normal, and the foals general attitude was improved. Subsequent feedings of milk replacer were given by bottle. A concentration immunoassay testa of plasma immunoglobulin concentration indicated adequate passive transfer (> 800 mg/dl), and results of a CBC were normal other than mild neutrophilia with a regenerative left shift. The foal was treated with procaine penicillin G (22,000 U/kg [10,000 U/lb], IM, q 12 h) and trimethoprim-sulfamethoxazole (30 mg/kg [13.6 mg/lb], PO, q 12 h) prophylactically for 7 days. A third transfusion consisting of 360 ml of the mares washed RBC in 400 ml of saline solution was given 36 hours after parturition because of the value of the filly and the availability of the RBC. The foals PCV after this transfusion was 23%, and the foal was allowed to resume nursing the mare at this time. The foal was bright and responsive but would show signs of exercise intolerance when stressed. Follow-up hematologic and serum biochemical testing was done periodically for the first 17 days after parturition. Serum total bilirubin concentration peaked on day 3 at 16.1 mg/dl (reference range, 0.5 to 3.9 mg/dl) and was normal on day 17. Packed cell volume was approximately 20% from day 2 through day 6 but decreased to 14% on day 9. After day 14, the PCV increased slowly and was within reference limits1 (30 to 49%) by day 28. Results of follow-up CBC and serum biochemical testing, in conjunction with the foals clinical appearance, indicated that it did not have any serious concurrent infections or systemic abnormalities. Follow-up physical examinations revealed progressive resolution of the icterus and increased tolerance to exercise. The foal recovered without complications. Blood typing analysis was done on the mare and the foals born during 1987, 1991, 1992, and 1994. All of these foals had different sires. All 4 foals were positive for the Qc, Db, and Dq antigens, and the 3 most recently born foals were also positive for the Ua antigen. The mare was negative for Qc, Db, Dq, and Ua
Scientific Reports: Clinical Report 79

' Neonatal isoerythrolysis is caused by destruction of neonatal erythrocytes by maternal alloantibodies in colostrum that are ingested and passively absorbed from the gastrointestinal tract. ' Approximately 90% of all cases of neonatal isoerythrolysis in horses are attributable to the Aa or Qa antigen; however, other antigens may be implicated.

EQUINE

n 1992, a multiparous 13-year-old Thoroughbred mare and its 48-hour-old colt were examined because of possible neonatal isoerythrolysis (NI). The mare had delivered the colt without incident; however, the owners were concerned that the mare did not have a sufficient quantity of colostrum. Therefore, on at least 2 occasions during the first 24 hours after parturition, the foal was given additional colostrum collected from other multiparous mares on the farm and stored frozen for up to 1 year. Within 48 hours of parturition, the foal developed clinical signs consistent with NI. Supportive treatment was administered, and the foal recovered without requiring a transfusion. According to the owners, the mare had delivered foals without incident during 1987 and 1991. The mare was barren during 1993, but in 1994 delivered a filly after approximately 337 days of gestation. The delivery was characterized by premature placental separation; delivery was promptly assisted, and no other complications were encountered. The foal weighed approximately 45 kg (99 lb) at birth and nursed vigorously within 2.5 hours after parturition. During a routine veterinary examination 12 hours after parturition, the foal was found to be tachypneic, icteric, and lethargic. Rectal temperature was 38.6 C (101.5 F), pulse rate was 160 beats/min, and respiratory rate was 22 breaths/min. The PCV was 10%. A diagnosis of NI was made on the basis of the past history of the mare, the physical examination findings, and the profound anemia. The foal was given a commercially available equine milk replacer, to which mineral oil had been added, and was then muzzled. Because of the foals unstable condition, the decision was made to manage the foal on-site. Lactated Ringers solution (1 ml/kg of body weight per hour [0.45 mg/lb/h]) was administered IV while preparations were made for a transfusion. Approximately 15 hours after parturition, blood was collected from the mare and submitted to a local human hospital for separation and washing of the
From the Department of Clinical Sciences, College of Veterinary Medicine and Biomedical Sciences, Colorado State University, Fort Collins, CO 80523. The author thanks Dr. Ann Bowling for technical assistance. JAVMA, Vol 219, No. 1, July 1, 2001

Table 1Results of blood typing analysis of a Thoroughbred mare and its 4 foals
Mare or foal Mare Foal Year of birth NA 1987 1991 1992 1994 Blood group A adf adf adf adf adf C a a a a a D cdgklm bcdklmq bcgmpq bcgmpq bcdklmq K P b Q c c abc U a a a

All foals were sired by different stallions; foals born during 1992 and 1994 developed clinical signs of neonatal isoerythrolysis. NA = Not applicable.

antigens (Table 1). Samples of the mares serum had been tested for titers of alloantibodies to all known blood group factors on 4 occasions by the blood typing genetics laboratory at the University of California at Davis. Two weeks after foaling in 1992, the mare was found to have high titers (> 1:16) of alloantibodies against and strong reactions to the Qc and Db antigens. Three weeks prior to foaling in 1994, the mare did not have detectable alloantibodies against Qc but had a high titer of alloantibodies against and a strong reaction to Db. Twenty-four hours after foaling in 1994, the mare had a high titer of alloantibodies (> 1:64) to Qc and had a 4-fold increase (> 1:256) in alloantibody titer against and a strong reaction to Db. Four months after foaling, titers of alloantibodies to Qc and Db had fallen (> 1:16), and the reaction to Qc was weak; however, the reaction to Db was still strong, despite the lower titer. A diagnosis of NI secondary to Qc and Db alloantibodies was made on the basis of results of blood typing. Neonatal isoerythrolysis is caused by destruction of neonatal erythrocytes by maternal alloantibodies that are ingested in colostrum and passively absorbed from the gastrointestinal tract. Once in the blood, alloantibodies attach to the neonatal erythrocytes and precipitate a hemolytic crisis through direct cell lysis, agglutination, or both.2-5 Neonatal isoerythrolysis is primarily seen in foals born to multiparous mares, and clinical signs are typically apparent between 8 and 96 hours after parturition.6 Alloantibodies can develop only when a mare is bred to a stallion that possesses an erythrocyte antigen that the mare does not share and the foal inherits that antigen. Alloantibodies are produced by the mare when she is exposed to the antigen through small transplacental hemorrhages that occur late in gestation or during parturition. Mares may also be sensitized through transfusion or immunization with products derived from blood or as a result of placental abnormalities during gestation.7 Because this initial immunization typically results in only low titers of alloantibodies and typically occurs late in gestation or during parturition, the chances that the foal produced during that pregnancy will develop clinical signs of NI are low. However, foals produced during subsequent pregnancies that have the same erythrocyte antigen are much more likely to develop NI. During the late stages of gestation, the mares antibody titers, including the titer of the antierythrocyte alloantibody, increase, and large quantities of antibodies are concentrated in the colostrum. When the foal ingests this colostrum, the
80 Scientific Reports: Clinical Report

antibodies are passively absorbed. Once in the blood, the alloantibodies act as agglutinins or lysins, and erythrocytes to which the alloantibodies have attached are cleared by the reticuloendothelial systems of the spleen and liver. In severe cases, death ensues as a result of hypoxia or disseminated intravascular coagulation. In previous reports and research surveys,8,b the antigens associated with NI have primarily been Aa and Qa, and approximately 90% of all cases of NI in horses are attributable to the Aa or Qa antigen. Other antigens that have been implicated in NI include Ab, Qrs, Qb, Qc, Dc, Da, Ka, Pa, and Ua.9-10,b On the other hand, the anti-Ca alloantibody may help protect against NI.11 Fourteen percent of all mares are considered to have blood types conducive to the production of alloantibodies against antigens implicated in NI.2 However, this tendency to produce alloantibodies varies by breed.7 Approximately 10% of Thoroughbred mares have been found to have some alloantibodies,8 but only a small percentage of these mares become alloimmunized and have a subsequent pregnancy with an anamnestic response strong enough and directed against the appropriate antigenic factors to result in NI.7 The overall incidence of clinical NI has been estimated to be 1% in Thoroughbreds and 2% in Standardbreds; however, subclinical NI may and probably does occur.7 The cause of the initial alloimmunization in the mare described in the present report was not known, but the mare was presumed to have been alloimmunized during previous pregnancies. The mare did not have any history of receiving any transfusions or of having been vaccinated with products containing blood or tissue. Possible causes of anemia, other than NI, in neonatal foals include blood loss from iatrogenic or obstetrical causes; perinatal hemorrhage seen as gastrointestinal tract, intra-abdominal, intrathoracic, or soft tissue hemorrhage secondary to trauma; thrombocytopenia; and hereditary bleeding disorders.12 With these disorders, total bilirubin concentration is often within reference limits initially, and an increase in bilirubin concentration is most commonly a result of hemolytic anemia. Causes of hemolytic anemia in foals include NI and acquired hemolysis secondary to snake venom, infection, and disseminated intravascular coagulation.12 Neonatal isoerythrolysis in the 1994 foal described in the present report was diagnosed on the basis of results of clinical examination, laboratory testing, and the history of NI in the 1992 foal. To the authors knowledge, a report of NI in a foal involving the Qc or Db antigen has not been published previously. The 1992 foal, which had clinical signs consistent with NI, was negative for the Qa antigen, as was the mare. Therefore, NI in this foal could not have been caused by alloantibodies to Qa. The 1994 foal was positive for the Qa antigen, but the mare did not have any detectable alloantibodies against Qa when tested 2 weeks after parturition in 1992 and when tested 3 weeks before and 2 days and 4 months after parturition in 1994. Because the 1987, 1991, and 1992 foals were all negative for the Qa antigen, it seems likely that the mare had not been previously exposed to this antigen, making it unlikely that clinical NI in the 1994 foal was
JAVMA, Vol 219, No. 1, July 1, 2001

EQUINE

a result of alloantibodies against Qa. Similarly, the mare did not have alloantibodies against Ua and did not react to Dp or Dq antigens. Therefore, these antigens were ruled out as possible causes of NI in the 1994 foal. The only D antigen that the mare reacted against was Db, and that reaction was characterized by strong agglutination and an increase in titer at the time of parturition and a subsequent decrease. The in vitro classification of lytic or agglutinating activity does not necessarily reflect in vivo activity of the antibody; however, the Db antigen has not previously been implicated in NI because of its agglutinating behavior.5 The Q family of antigens has been implicated in NI, and in this mare the anti-Qc titer rose from none 3 weeks prior to parturition to > 1:64 24 hours after parturition and fell to > 1:16 by 4 months after parturition in 1994. In 1992, the mares anti-Qc titer 2 weeks after parturition was > 1:16, and the mare reacted strongly to Qc; the 1992 foal was also positive for the Qc antigen. It is unknown whether the foal born in 1991 had any evidence, clinical or subclinical, of NI. However, one may hypothesize that 3 sequential pregnancies after the first in 1987 resulting in foals positive for the Db and Qc antigens may have increased the mares sensitivity to these antigens and led to clinical signs of NI in the 1994 foal. However, diagnostic testing was not helpful in determining whether the Db or the Qc antigen was more responsible for NI in the 1994 foal than the other, or whether both were necessary for clinical disease. The mares strong agglutinating reaction against the Db antigen and lytic reaction against the Qc antigen may have cumulatively precipitated the hemolytic crisis. Peri-parturient testing of mares for alloantibodies to Aa and Qa has proved successful in managing mares susceptible to producing foals with NI. However, the mare described in the present report was tested for alloantibodies at the recommended time, 2 weeks before the anticipated due date in 1994,2 and antibody titers at that time did not indicate that the coming foal could not nurse the mare. Thus, peri-parturient testing

may not be foolproof in all cases. In suspect situations, a dam-side jaundice foal agglutination test, using either the mares serum13 or colostrum,9 may prove useful.
a b

Cite, Idexx, Portland, Me. Cox LA. A survey of equine red blood cell antibody frequencies and their relevance to neonatal isoerythrolysis. Clinical sciences and genetics. MS Thesis, Genetics Laboratory, University of California, Davis, Calif, 1984.

References
1. Leadon DP Clinical pathology data. In: Robinson NE, ed. . Current therapy in equine medicine. 3rd ed. Philadelphia: WB Saunders Co, 1992;822828. 2. Becht JL. Neonatal isoerythrolysis in the foal, part 1: background, blood group antigens and pathogenesis. Compend Contin Educ Pract Vet 1983;5:S591S596. 3. Scott AM, Jeffcott LB. Haemolytic disease of the newborn foal. Vet Rec 1978;103:7174. 4. Trommershausen-Smith A, Stormont C, Suzuki Y. Alloantibodies: their role in equine neonatal isoerythrolysis, in Proceedings. 1st Int Symp Equine Hematol 1975;349355. 5. Witham CL, Carlson GP Bowling AT. Neonatal isoerythrol, ysis in foals: management and prevention. Can Vet J 1984;11:2123. 6. Kahn W, Vaala WE, Palmer J. Neonatal isoerythrolysis in newborn foals. Tierarztl Praxis 1991;19:521529. 7. McClure JJ, Parish SM. Diseases caused by allogeneic incompatibilities. In: Smith BP, ed. Large animal internal medicine. 2nd ed. St Louis: Mosby, 1996;18621873. 8. Bailey E. Prevalence of anti-red blood cell antibodies in the serum and colostrum of mares and its relationship to neonatal isoerythrolysis. Am J Vet Res 1982;43:19171920. 9. Bailey E, Conboy HS, McCarthy PF Neonatal isoerythroly. sis of foals; an update on testing, in Proceedings. Am Assoc Equine Pract 1987;341349. 10. Zaruby JF Hearn P Colling D. Neonatal isoerythrolysis in a , , foal involving anti-Pa alloantibody. Equine Vet J 1992;24:7173. 11. Bailey E, Albright DG, Henney PJ. Equine neonatal isoerythrolysis: evidence for prevention by maternal antibodies to the Ca blood group antigen. Am J Vet Res 1988;49:12181222. 12. Vaala WE. Neonatal anemia. In: Koterba AM, ed. Equine clinical neonatology. Philadelphia: Lea & Febiger, 1990;571588. 13. Becht JL, Page EH, Morter RL, et al. Evaluation of a series of testing procedures to predict neonatal isoerythrolysis in the foal. Cornell Vet 1983;73:390402. EQUINE

JAVMA, Vol 219, No. 1, July 1, 2001

Scientific Reports: Clinical Report

81