Pathology in Practice

Bianca R. Pfisterer Cummings School of Veterinary Medicine, Tufts University, North Grafton, MA 01536 (Pfisterer)

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Kara N. Corps Department of Population Health and Pathobiology, North Carolina State University, Raleigh, NC 27607

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Samuel H. Jennings Department of Population Health and Pathobiology, North Carolina State University, Raleigh, NC 27607

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History

A 14-year-old 522-kg (1,148.4-lb) Quarter Horse gelding was referred to North Carolina State University Equine Emergency Services because of signs of pain of 1 week's duration that were attributable to a nonresolving, malodorous laceration over the lateral aspect of the right thigh. The lesion had a blood-tinged discharge and crepitation. The day prior, the referring veterinarian removed an approximately 7-cm-long, wooden foreign body from the semimembranosus and semitendinosus muscles. Prior treatment included administration of ceftiofur, fluid therapy, dimethyl sulfoxide, and NSAIDs.

Figure 1—
Figure 1—

Photograph obtained during necropsy of a 14-year-old Quarter Horse gelding that was evaluated because of swelling and a laceration associated with the semitendinosus and semimembranosus muscles of the right hind limb. In this view, a transverse section of the caudal aspect of the right thigh musculature at the level of the clinically identified injury is shown. There is extensive dark red–black discoloration of the muscle caused by necrosis and hemorrhage with marked expansion of the neighboring fascia caused by edema.

Citation: Journal of the American Veterinary Medical Association 254, 6; 10.2460/javma.254.6.681

Clinical and Gross Findings

On physical examination, the horse had signs of depression and was hypothermic, tachycardic, and tachypneic. The horse had a pain score1 of 30/36; therefore, butorphanol tartrate, detomidine hydrochloride, and acepromazine maleate were administered. Extending through the skin and into the right semimembranosus and semitendinosus muscles was an approximately 10-cm-long laceration; ultrasonography confirmed marked swelling in the area. Abdominocentesis yielded blood-tinged exudate with 6,200 WBCs/μL (< 5,000 WBCs/μL being considered normal), BUN concentration of 37 mg/dL, and creatinine concentration of 5.2 mg/dL. A CBC and serum biochemical analysis revealed polycythemia (PCV, 55%; reference interval, 31% to 48%) with hypoproteinemia (4.2 g/dL; reference interval, 5.9 to 8 g/dL), azotemia (BUN concentration, 30 mg/dL [reference interval, 7 to 25 mg/dL]; creatinine concentration, 4.5 mg/dL [reference interval, 1 to 1.7 mg/dL]), hyperglycemia (176 mg/dL; reference interval, 73 to 113 mg/dL), hyperkalemia (6.9 mmol/L; reference interval, 2.9 to 4.8 mmol/L), hypocalcemia (9.6 mg/dL; reference interval, 11.3 to 13.4 mg/dL), and high aspartate aminotransferase activity (966 U/L; reference interval, 202 to 339 U/L). Fluid therapy was started, and another dose of butorphanol tartrate was given. Shortly thereafter, the horse became laterally recumbent. Euthanasia by IV administration of pentobarbital sodium and phenytoin sodium was performed because of the horse's poor prognosis and nonresolving signs of pain. The carcass was submitted for necropsy.

Gross examination revealed marked, subcutaneous edema of the right hind limb, inguinal region, caudoventral portion of the abdomen, and prepuce. A 10 × 4-cm penetrating laceration at the junction of the right semimembranosus and semitendinosus muscles, equidistant from the tuber ischia and tuber calcanei, extended 15 cm deep into the underlying musculature and soft tissues. The surrounding muscle and connective tissue were markedly friable, edematous, and extensively dark red to purple with multifocal black discolorations (Figure 1). The surrounding vastus medialis, gracilis, and sartorius muscles had extensive hemorrhage with occasional pockets of emphysema.

Formulate differential diagnoses from the history, clinical findings, and Figure 1—then turn the page

Cytologic, Microbiological, and Histopathologic Findings

Cytologic examination of samples of affected muscle revealed numerous long bacterial rods with clear discrete spores consistent with Clostridium spp. Fresh muscle tissue samples were submitted to the Rollins Animal Disease Diagnostic Laboratory for clostridial fluorescent antibody testing, and the tissue samples were positive for Clostridium septicum. Skeletal muscle from the right hind limb and adjacent adipose tissue were fixed in neutral-buffered 10% formalin, routinely processed, and stained with H&E stain for histologic examination. The skeletal muscle and adjacent adipose tissue had large zones of coagulative to liquefactive necrosis, with hemorrhage, fibrin, and edema (Figure 2). Necrosis was characterized by loss of cross striations, sarcoplasmic hypereosinophilia, and nuclear pyknosis and karyorrhexis. Multifocally, small to moderate numbers of degenerate neutrophils and fewer macrophages infiltrated the affected skeletal and adipose tissue. Occasionally, clusters of large bacilli with a few clear spores were present. Gas production was supported by scattered large, round clear spaces within the tissues. Some blood vessels within the lesions contained fibrin thrombi or had evidence of fibrinoid necrosis.

Morphologic Diagnosis and Case Summary

Morphologic diagnosis and case summary: severe, extensive, necrohemorrhagic and emphysematous myositis and steatitis with intralesional spore-forming bacilli (consistent with malignant edema attributable to C septicum infection) in a horse.

Comments

In the case described in the present report, the gross and histologic findings were consistent with a diagnosis of malignant edema, also known as clostridial myositis. The horse's polycythemia was interpreted as relative polycythemia resulting from a combination of dehydration, third-space migration of fluid as edema, and potentially a contribution by epinephrine-induced splenic contraction. Prerenal factors were likely a major contributor to the azotemia, although sepsis-related renal dysfunction may also have had a role. The horse's hyperkalemia and high aspartate aminotransferase activity were suspected to be primarily caused by muscle necrosis, although sepsis-related renal dysfunction could have contributed to the hyperkalemia. The horse's hypocalcemia may also have been multifactorial. The horse's serum albumin concentration was within the reference interval but at the lower limit (2.3 g/dL [reference range, 2.2 to 3.7 g/dL]), which may have partially contributed to the reduction in the measured serum total calcium concentration. Serum phosphorus concentration was not measured, so it was unknown whether hyperphosphatemia from muscle injury or decreased glomerular filtration (prerenal or renal causes) may have contributed to the horse's hypocalcemia. The mildly high cell count in the fluid sample obtained via abdominocentesis may have been related to systemic inflammation or early direct extension of inflammation from the limb into the peritoneum.

Figure 2—
Figure 2—

Photomicrographs of skeletal muscle sections from the horse in Figure 1. A—The myofibers are diffusely hypereosinophilic and lack nuclei. Frequently, individual myofibers and fascicles are variably separated by clear space (edema or gas [asterisk]), eosinophilic fluid (edema), and hemorrhage. Basophilic aggregates of bacteria are just discernible (dagger). H&E stain; bar = 500 μm. B—At a higher magnification, loss of cross striations with sarcoplasmic fragmentation in addition to nuclear loss by myofibers is evident. A blood vessel has undergone fibrinoid vascular necrosis with disruption of its wall by fibrin and nuclear debris (asterisk). A few colonies of bacteria are visible (dagger). H&E stain; bar = 100 μm. C—The bacterial colonies are composed of large, gram-positive bacilli. Gram stain; bar = 50 μm.

Citation: Journal of the American Veterinary Medical Association 254, 6; 10.2460/javma.254.6.681

Clostridium spp are gram-positive, spore-forming, anaerobic bacteria that are ubiquitous in the environment and gastrointestinal tract of animals.2–4 In aerobic environments, sporulation of the bacteria results in dormant spores (thereby promoting survival), whereas in anaerobic environments, spore germination occurs and results in vegetative cells that can proliferate and produce exotoxins.4 Various diseases (clostridial myositis, blackleg, black disease, so-called big head of sheep and goats, bacillary hemoglobinuria, enterotoxemia, tetanus, and botulism) are caused by infections with bacteria in the genus Clostridium; the development of each disease is dependent on the species of Clostridium, the species of animal affected, and the inoculation site.5

Clostridial myositis in sheep, cows, humans, and horses has been reported.6 Numerous clostridial species, most commonly Clostridium perfringens type A and C septicum, are capable of infecting and causing myositis in horses.2,3,6 Each species of Clostridium produces different exotoxins, of which α toxin (phospholipase c) is the most important in the pathogenesis of clostridial myositis resulting from infection with C perfringens type A or C septicum.2,4,6,7 The α toxin inhibits leukocyte infiltration, promotes cell membrane damage and hemolysis, and increases vascular permeability.8,9

Horses typically develop clostridial myositis through soft tissue injuries including surgical sites, lacerations, or IM injections.2 Most cases have been associated with IM injections.2,6 After tissues are injured, local ischemia develops and generates an anaerobic environment that propagates spore germination.5 Within 6 to 72 hours, the clostridial toxins cause the horse to become febrile with localized edema and signs of pain, heat, and crepitation that rapidly progress to a firm, cool, discolored lesion with a foul odor.6,8 The widespread cellular and vascular damage from the systemic toxemia leads to cardiovascular shock, multiorgan dysfunction, and frequently death within 72 hours after inoculation, if untreated.8 In addition to environmental inoculation, Clostridium histolyticum, Clostridium sporogenes, and C perfringens can be found in healthy skeletal muscle tissue, but it is not known whether the presence of such organisms has a role in the development of clostridial myositis.3

Survival of horses infected with Clostridium spp requires early and aggressive treatment.2,6,9 The best approach to treatment includes IV administration of high dosages of appropriate antimicrobials, provision of supportive care, and surgical debridement or fasciotomy to decrease the amount of necrotic tissue and aerate the muscle.6,9 The choice of antimicrobials used varies greatly, with penicillin G potassium being frequently used in combination with other antimicrobials such as gentamicin, amikacin, or metronidazole.9 The infective Clostridium spp appears to affect survival rate among horses with clostridial myositis; horses infected with C perfringens have a better survival rate than that of horses infected with C septicum or multiple clostridial species.6 In 1 retrospective study6 of 37 horses, those with C perfringens infection alone had a survival rate of 81% and those with C septicum infection alone had a survival rate of 50%, which are both much higher survival rates than those previously reported.2

References

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  • 8. Songer JG. Histotoxic clostridia. In: Gyles CL, Prescott JF, Songer G, et al, eds. Pathogenesis of bacterial infections in animals. 4th ed. Ames, Iowa: Blackwell Publishing, 2010;203209.

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  • 9. Adam EN, Southwood LL. Surgical and traumatic wound infections, cellulitis, and myositis in horses. Vet Clin North Am Equine Pract 2006;22:335361.

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