Massive intravascular hemolysis secondary to Clostridium perfringens septicemia is rare but often fatal. We report a case of a fatal clostridial hemolytic complication in a 71-year-old woman with probable refractory anemia. The patient was admitted to the emergency room due to a comatose mental state and a high fever. Laboratory analysis showed massive hemolysis. She died from severe anemia two hours after admission. The next day, blood cultures grew gram positive cocci and boxcarshaped gram positive rods, which were identified as coagulase-negative staphylococci and C. perfringens, respectively.

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Massive intravascular hemolysis is a rare but often fatal complication of Clostridium perfringens bacteremia<1>. This feature is reported in the groups of patients who have undergone abortion, childbirth, or trauma, who are immunocompromised because of diseases such as diabetes and malignancy including leukemias, and who are apparently healthy but are found at autopsy to have a portal of entry for the bacteria, such as liver or spleen abscess, endocarditis, cholecystitis, gastrointestinal arteriovenous malformations, or pleural effusions<2-4>. But we could find only one case of such complication in a patient with refractory anemia<5>. We report a case of a 71-year-old woman with probable refractory anemia who presented with massive intravascular hemolysis and died from severe anemia 2 hr after admission.

A 71 year-old-woman with jaundice and coma of recent onset was admitted to emergency room. She had been diagnosed of probable refractory anemia about 6 months before, when her white blood cell count (WBC) was 1.28×109/L (neutrophils 22%, lymphocytes 72%, and blasts 2%), hemoglobin (Hb) 9.0 g/dL, hematocrit 26.8%, mean corpuscular volume (MCV) 100.9 fL, mean corpuscular hemoglobin (MCH) 33.9 pg, and platelets 42×109/L. Bone marrow aspiration and biopsy were not attained because of her refusal. She was followed up for months, and received on the average 2 units of packed red cells a month. On the present admission, she was icteric, and her lips were cyanotic. The blood pressure was 106/49 mm Hg, heart rate 120/min, and body temperature 40.5℃. One blood culture set was drawn immediately and laboratory analysis was performed. Her WBC was 6.21×109/L (neutrophils 24%, lymphocytes 50%, and atypical lymphocytes 24%), Hb 2.2 g/dL, hematocrit 1.4%, MCV 84 fL, and platelets 28×109/L. The serum and plasma was red-colored. Other laboratory results are shown in Table 1. Peripheral blood smear demonstrated a markedly decreased number of the red cells (not more than 10 red cells per high power field <×400>) with spherocytosis, and occasional extracellular microorganisms (Fig. 1), but gram stain was not performed with blood smears. The patient was transfused 2 units of packed red cells, but thereafter became asystolic, and died 2 hours after admission. No additional laboratory test or postmortem examination was performed. The next day blood culture grew bacteria.
Fig. 1Peripheral blood smear showed rare intact red cells with spherocytes, and occasional rod-shaped microorganisms (×1,000). But it was not determined whether the microorganisms were pathogens or contaminants.
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Table 1Results of laboratory tests except complete blood count
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Within five minutes of the patient"s admission, the blood drawn from a peripheral vein was infused into a blood culture set consisting of a BACTEC Aerobic/F culture bottle (Becton Dickinson, Sparks, Maryland, USA) and a BacT/Alert anaerobic culture bottle (bioMerieux, Durham, North Carolina, USA). Culture bottles were incubated in BACTEC 9240 system (Becton Dickinson) and BacT/Alert 3D system (bioMerieux). The next day, bacteria grew in the culture bottles. The bacteria were gram positive cocci and gram positive rods. Gram positive cocci were subcultured onto blood agar and identified as coagulase-negative staphylococci. Gram positive rods were boxcarshaped and were subcultured onto brucella vitamin K1 blood agar (BRBA) in Anaerobic Pouch System (Oxoid Ltd., Basingstoke, Hampshire, England). Incubated for 48 hr, colonies showed irregular edge and double zone of hemolysis (Fig. 2). The rods were identified as C. perfringens (profile code: 4501404030; %id: 99.9; and T index: 0.42) by a commercial identification kit, API ID 32 A (bioMerieux, Marcy-l"Etoile, France).
Fig. 2Colonies grown on blood agar plate showed double zone hemolysis, which comprised an inner zone of complete hemolysis and an outer zone of incomplete hemolysis.
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Clostridia account for only about 10% to 12% of the anaerobic bacteria isolated from properly selected and collected clinical specimens<6>, and less than 3% of all blood cultures<7>. C. perfringens is the most common clinical isolate of Clostridium<2, 6, 7>, but as many as 50% of isolates from the blood are assessed to be contaminants<7>. C. perfringens is nonmotile and generally has a distinctive box-car appearance on gram stain of clinical material or subcultures<7>, It is commonly found among the normal flora of the gastrointestinal tract, therefore, the vast majority of infections are of endogenous origin<3, 6>. Usually the bacteria gain entrance to the body through penetrating wounds or mucosal defects in the gastrointestinal, genitourinary, or hepatobiliary tract<1>, and break the gut barrier with ease in patients with leukemia, malignancy, or hepatic cirrhosis, especially when there is a loss of integrity of the bowel<3>. In neutropenic patients including our case, the organisms can readily enter the blood stream, even in the absence of obvious gut pathology<3>. But we could not find a report that describes the relationship between refractory anemia and C. perfringens-induced massive hemolysis.

The clinical spectrum of C. perfringens bacteremia ranges from a positive culture without associated symptoms to the syndrome of rapid shock and death<2>. C. perfringens produces one or more major lethal toxins (alpha, beta, epsilon, iota toxins, etc). Among these, alpha toxin (alpha-lecithinase, phospholipase C) is the most important toxin and produced by all types of C. perfringens<8>. Intravascular hemolysis is mediated by alpha toxin, which hydrolyzes sphingomyelin and lecithin to phosphoryl choline and diglyceride and lyses red blood cells, white blood cells, platelets, endothelial cells, and the plasma membranes of muscle cells<1, 3>. Automated blood counts might show disproportionately high hemoglobin levels in comparison with hematocrit, artificially increased MCH, and very low MCV<9>. In our case, the MCH and mean corpuscular hemoglobin concentration (MCHC) were spuriously increased to 136.5 pg and 162.5 g/dL, respectively, which is probably due to free plasma hemoglobin.

The definite treatment for clostridial septicemia is administration of antimicrobial agents and aggressive surgical debridement of any potential source of infection<1, 2>. High-dose penicillin, preferably by continuous intravascular infusion, was known as the drug of choice, but several studies have reported an increased resistance of C. perfringens to various antibiotics including penicillin G<2-4, 10>. Clindamycin, metronidazole, second or third generation cephalosporins, chloramphenicol, and imipenem will be alternative agents<1, 3, 4, 10>. Exchange blood transfusions with packed or washed red blood cells, and hyperbaric oxygen has also been tried<1, 4>. Despite all therapeutic efforts, the prognosis is poor, with reported mortality ranging from 35% to almost 100%<1, 8, 10, 11>. In our case, no empiric antimicrobials were started in spite of high fever. But even if any antibiotic treatment was started, the patient could hardly survive due to the rapid progression of intravascular hemolysis. Postmortem examination was not performed, and the infection source was not identified.

In Korea, there has been no reported case of massive intravascular hemolysis caused by C. perfringens infection, except for a case of liver abscess with sepsis and suspicious intravascular hemolysis by C. perfringens<12>. In our case, though only one blood culture set was drawn, fever, massive intravascular hemolysis, and rapid growth of bacteria are very well consistent with C. perfringens sepsis. In addition, there was no evidence of known causes of massive intravascular hemolysis, including malaria, bartonellosis, babesiosis, hemolytic uremic syndrome, blood transfusion, snake venoms, or extensive acute burns.

Gutierrez et al.<13> postulated that clinical physicians must suspect clostridial septicemia in a patient who presents with fever, jaundice, and intravascular hemolysis, especially with underlying malignant conditions or softtissue involvement. Although C. perfringens-induced hemolytic anemia is rare, early recognition of C. perfringens is very important because only a prompt antibiotic therapy before the anemia became life-threatening can rescue patients from an otherwise rapidly fatal outcome<9>.

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13. Gutierrez A, Florencio R, Ezpeleta C, Cisterna R, Martinez M. Fatal intravascular hemolysis in a patient with Clostridium perfringens septicemia. Clin Infect Dis 1995;20:1064–1065.
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