Heinz Body Hemolytic Anemia With Eccentrocytosis From Ingestion of Chinese Chive (Allium tuberosum) and Garlic (Allium sativum) in a Dog

Introduction

In dogs and cats, onion (Allium cepa) is known to be oxidatively toxic to erythrocytes, resulting in hemolytic anemia.¹² Ingestion of garlic (Allium sativum) can also experimentally induce hemolysis in dogs as a result of oxidative damage to erythrocytes.³ Other members of the genus Allium (A.) are not known to cause hemolysis in dogs and cats, although wild onion (A. validum and A. canadense) and wild garlic (A. ursinum) are reported to cause hemolytic anemia in ruminants and horses.^4–6

Chinese chive (A. tuberosum) is a strong-flavored herb of commercial importance in East Asia, especially in Japan, Korea, and China. It has edible foliage leaves, but no bulbs.⁷ The external appearance of this plant resembles that of A. ampeloprasum, which includes several distinct crop types (i.e., leek, kurrat, great-headed garlic, pearl onion) from Europe and the United States. Chinese chive has not been reported to be toxic to domestic animals.

The following case report describes an adult dog that developed severe hemolytic anemia after ingesting Chinese food containing Chinese chive and garlic. To the authors’ knowledge, this is the first report of naturally occurring hemolytic anemia in a dog caused by Chinese chive and/or garlic.

Case Report

A 4-year-old, 7.2-kg, intact male miniature schnauzer was presented to a private animal hospital for anorexia unassociated with vomiting and diarrhea, starting the previous night. The owner reported the dog had ingested some Chinese steamed dumplings 2 days before and that the urine was dark in color for 1 day. The steamed dumpling was composed of minced pork and vegetables wrapped in a small pancake, and it contained Chinese chive and garlic. The amount of these Allium plants ingested by the dog was unknown. Initial physical examination revealed a mildly depressed dog with a rectal temperature of 102.4°F (39.1°C) and slightly pale, dark mucous membranes. Collected venous blood was dark brown in color.

A complete blood count at admission (day 2 postingestion) revealed a mild anemia (hematocrit [Hct] of 33%; reference range, 36% to 54%), reddish plasma from hemolysis, and a high mean corpuscular hemoglobin concentration (MCHC) of 36.5% (reference range, 30.5% to 36.1%) [Figure 1]. Moderate leukocytosis (17,500/μL; reference range, 5100 to 10,700/μL) with a neutrophilia (11,200/μL; reference range, 1400 to 8100/μL) and a left shift (bands 1663/μL; reference range, 50 to 900/μL) were also noted [Figure 2]. Analysis of a stained^a blood smear revealed many eccentrocytes with hemoglobin that appeared dense and contracted to one side of the cell, with a pale area on the other side and projections on the cell surface that looked like Heinz bodies [Figure 3]. The percentage of eccentrocytes on day 2 was 37.5% of all erythrocytes [Figure 4]. Results of blood chemistry analyses at initial presentation were within normal limits.

Based on these hematological findings, oxidative damage to erythrocytes was suspected from the ingestion of Chinese chive and garlic.

Initial therapy at the private animal hospital included subcutaneous lactated Ringer’s solution with B-complex vitamins to promote diuresis and prevent formation of hemoglobin casts and tubular nephrosis; famotidine^b (0.7 mg/kg subcutaneously [SC]) to prevent gastric ulceration; and orbifloxacin^c (5 mg/kg SC) to prevent secondary infections. In addition, human recombinant erythropoietin (epoetin alpha,^d 50 IU/kg SC) was administered to accelerate regeneration of erythroid cells. On day 3 postingestion, the Hct had decreased to 23%, and the MCHC had increased to 37.4% secondary to hemolysis [Figure 1].

On day 4 postingestion, the dog was presented to the Veterinary Teaching Hospital at Hokkaido University for evaluation of progressive anemia. Pale mucous membranes were observed on physical examination, and collected venous blood appeared dark brown. Hemolysis was not observed in the serum, and the MCHC had returned to normal (34.2%). The Hct reached a nadir (19%) on day 4 postingestion [Figure 1]. At the same time, the percentage of eccentrocytes remained high (21.6%), and erythrocytes containing Heinz bodies accounted for 44.3% of all the erythrocytes [Figure 4]. Methemoglobin concentration was measured using the method reported by Hegesh et al. and was found to be high (2.1%) [Figure 5] compared to the reference range (<1%).⁸ Methemoglobinemia was thought to be the cause of the dark color of the blood. Persistent leukocytosis (16,400/μL) and a decrease in the platelet count (114,000/μL; reference range, 160,000 to 600,000/μL) were also observed [Figure 2].

The dog’s anorexia and slight depression were treated with lactated Ringer’s solution SC, supplemented with vitamin B-complex and vitamin C, oral administration of a concentrated high-calorie liquid diet,^e and an oral vitamin and mineral supplement.^f

The antioxidative status of the dog was examined using erythrocytes collected on day 4 to rule out a high susceptibility to oxidative stress as a genetic trait. The concentration of reduced glutathione and the activities of superoxide dismutase, catalase, glutathione peroxidase, glutathione reductase, reduced nicotinamide adenine dinucleotide methemoglobin reductase, glucose-6-dehydrogenase, and 6-phosphogluconate dehydrogenase were measured according to the methods described by Beutler.⁹ The reduced glutathione concentration (8.5 μmol/gHb; reference range, 6.1 to 13.6 μmol/gHb) [Figure 5] and all the enzyme activities were within reference ranges or slightly higher than their reference ranges (probably because of the high percentage of young erythrocytes in circulation). In addition, results of an osmotic fragility test and analysis of hemoglobin composition using high-performance liquid chromatography were also normal.¹⁰¹¹

On day 5 postingestion, the Hct value had increased (21%), indicating a regenerative response. The regeneration of erythrocytes also resulted in a higher mean cell volume (MCV, 83.7 fL; reference range, 57 to 66 fL) and a greater percentage of circulating reticulocytes (4.5%) [Figure 6]. The oral administration of vitamin E^g (tocopherol acetate; 7 mg/kg q 12 hours) was also initiated to prevent further oxidative damage to erythrocytes.

On day 9, the dog’s appetite and activity level began to recover. The erythrocyte regenerative response was also dramatic, with an MCV of 93.3 fL [Figure 1] and a reticulocyte count of 10.1% [Figure 6]. The Hct had increased to 25%, and the reduced glutathione concentration in erythrocytes was also increased (9.4 μmol/gHb) [Figure 5]. The reduced glutathione elevation was probably attributable to the release of young erythrocytes. The number of eccentrocytes (1.6%), the number of erythrocytes with Heinz bodies (5.7%), and the methemoglobin concentration (0.6%) were markedly decreased. The platelet count continued to decline until day 9 (82,000/μL), while the leukocyte count (10,400/μL) returned to normal [Figure 2].

Fluid therapy and the concentrated high-calorie liquid diet were withdrawn on day 9, but the vitamin E (7 mg/kg q 24 hours) and a commercial supplement^f of vitamins and minerals were continued until day 22. At day 22 postingestion, the anemia was improved and all other hematological data were almost normal. Hemolytic anemia has not recurred, and the dog has remained hematologically normal for several years.

Discussion

Heinz bodies arise from the denaturation of hemoglobin through a sequence of oxidative steps.¹²¹³ Methemoglobin forms when the iron moiety of heme protein is oxidized from the ferrous (II) to the ferric (III) state. Erythrocytes containing Heinz bodies are less deformable than normal erythrocytes.¹⁴ They tend to become sequestered in the reticuloendothelial system, where the Heinz bodies become pitted and the entire cell may be phagocytosed by macrophages.¹⁵ Eccentrocytes may form as a result of direct oxidative injury to erythrocyte membranes by adhesion of internal aspects of the cell membrane.^16–19 Eccentrocytes are rigid and spheroid with intrinsic membrane alterations that make them susceptible to entrapment and removal by the mononuclear phagocytic system.¹⁶ Heinz bodies and collapsed membranes of eccentrocytes become pitted in the reticuloendothelial system, and residual spheroid regions have increased osmotic fragility that may result in intravascular hemolysis. In the case reported here, eccentrocytosis, Heinz body formation, methemoglobinemia, and evidence of hemolysis were all observed. These hematological abnormalities were speculated to result from oxidative damage to erythrocytes caused by constituents in Chinese chive and/or garlic.

Eccentrocytosis has been reported in dogs given high doses of acetylphenylhydrazine, from prolonged administration of acetaminophen, and following the ingestion of dehydrated onions or hot-water extract of garlic.^3162021 In garlic-induced oxidative injury, the appearance of eccentrocytes may be the most prominent hematological feature, suggesting that the formation of eccentrocytes is a primary cause of garlic-induced hemolysis.³ In the case reported here, eccentrocytosis seemed to be a hallmark of the disease. The eccentrocytes were easily detected using any staining method. In contrast, the demonstration of Heinz bodies often required supravital staining using new methylene blue, brilliant green, etc., although they are generally visible using Romanowsky-type stains. Based on these findings, the detection of eccentrocytes may be useful in the diagnosis of Allium plant-induced oxidative injury.

One of the major flavor components of onions, n-propyl disulfide, is generally believed to be responsible for onion-induced hemolytic anemia.²² Three alk(en)yl thiosulfates (i.e., sodium n-propyl thiosulfate, sodium trans-1-propenyl thiosulfate, and sodium cis-1-propenyl thiosulfate) extracted from boiled onions are also thought to be causative agents of onion-induced hemolytic anemia.²³ In garlic-induced hemolysis, a novel alkenyl thiosulfate derivative, sodium 2-propenyl thiosulfate, has been isolated from boiled garlic and is considered to be one of the causative agents.²⁴ In addition, other volatile organosulfur compounds that oxidize canine erythrocytes have been isolated from an aqueous ethanol garlic extract, and the composite effects of these compounds on erythrocytes may be responsible for garlic-induced hemolysis.²⁵²⁶ Although no oxidants have been identified in Chinese chive, this plant is thought to contain oxidants similar to those in garlic, because Chinese chive and garlic possess similar flavor precursors specific for Allium plants (i.e., S-alk(en)yl cysteine sulfoxides).²⁷

Among Japanese and Korean purebred dogs (e.g., the Shiba and Jindo breeds), some individuals are particularly susceptible to onion-induced hemolytic anemia.²⁸²⁹ The high susceptibility of these dogs occurs from inherited high concentrations of erythrocyte reduced glutathione, which accelerates the oxidative damage produced by the sodium n-propyl thiosulfate and its derivatives found in onion and garlic.³⁰³¹ The dog reported here had a normal concentration of erythrocyte reduced glutathione, normal or slightly higher activities of antioxidative enzymes, and a normal composition of hemoglobin types, suggesting that the dog was genetically normal. It was therefore concluded that consumption of Chinese chive and garlic caused the hemolytic anemia experienced in this dog.

In the present case, human recombinant erythropoietin was used at a dosage of 50 IU/kg per day SC for 2 days in the early stages of the disease. This drug is generally administered for aplastic anemia associated with chronic renal failure, not for hemolytic anemia. However, the use of this drug in the initial stages of the anemia may have stimulated the production of erythrocytes earlier than the secretion of intrinsic erythropoietin, and this may have resulted in a more rapid resolution of the anemia. In addition, vitamin E was given at a dosage of 7 mg/kg q 12 to 24 hours from day 4, for about 3 weeks. The major physiological effect of vitamin E on erythrocytes is to act as a biological antioxidant protecting erythrocyte membranes from peroxidative damage.³² Vitamin E may be efficacious in the reduction of subsequent oxidative damage of erythrocyte membranes. Although no control dogs were utilized in this study, erythropoietin and vitamin E may have been beneficial in the dog’s recovery.

Conclusion

The ingestion of Chinese chive and garlic was followed by Heinz body hemolytic anemia with eccentrocytosis in a genetically normal adult dog. Results of hematological examinations, especially the detection of eccentrocytes, supported the diagnosis of Allium plant-induced hemolysis in the affected animal. This was the first report of naturally occurring hemolytic anemia in a dog associated with the ingestion of Chinese chive and/or garlic. Foods containing Chinese chive and garlic, as well as onions, should be avoided when feeding dogs.