INTRODUCTION

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Campylobacters are small, curved, motile, microaerophilic gram-negative rods. Campylobacter jejuni is the most common cause of bacterial diarrhea in the United States. However, C. jejuni bacteremia is uncommon; only about 0.4% of reported C. jejuni isolates originate from blood (1), and the bacteremia usually occurs in patients with underlying medical conditions or at the extremes of age (3). Likewise, disseminated infection by fungi of the order Mucorales, known as zygomycosis, is a rare infection and typically occurs in those immunocompromised by malignancy or renal disease (16). Here we report a case in which exertional heatstroke provided the setting for both disseminated zygomycosis and Campylobacter bacteremia. The zygomycete cultured was Rhizopus schipperae. This is only the second isolate of R. schipperae that has been described.

	CASE REPORT

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In June 1998, a 21-year-old Mexican woman crossed the United States border without documentation. She had a sore throat and cough prior to departure. She left San Luis Potosi, México, and rode a bus to Nuevo Laredo. With a group of men, she crossed the Rio Grande by boat and hiked into Dimmit County, Tex. In the morning, the woman had chills and diaphoresis. The immigrants drank from stock ponds and walked for several hours. The high temperature on this day was 105°F, and the woman became delirious and collapsed. She had convulsions and was taken to a local hospital. Initial vital signs were a rectal temperature of 107.1°F, a blood pressure of 71/44 mm Hg, a heart rate of 123 beats per minute, and a respiration rate of 30 breaths per minute. The patient was intubated and transferred to University Hospital in San Antonio, Tex. The assessment was exertional heatstroke with circulatory collapse, severe brain injury, shock liver, rhabdomyolysis, renal insufficiency, pancreatitis, and disseminated intravascular coagulation (DIC). Due to the probable exposure to enteric pathogens, levofloxacin was initiated on admission after blood and stool samples for cultures were obtained. The patient developed a lung infiltrate on day 3, and Enterobacter cloacae and a single colony of a Rhizopus species grew from tracheal aspirates. Blood and stool cultures grew C. jejuni. The antibiotic was changed to meropenem. On day 3, E. cloacae was also isolated from a blood culture.

On day 9, a 5-mm dark eschar was noted on her right nasal ala (Fig. 1A). Resection of the ala, lower lateral cartilage, and a portion of the nasal septum was performed until bleeding tissue was obtained (Fig. 1B). The nasal tissue showed nonseptate fungal hyphae (Fig. 1C). Amphotericin B (AMB) was started at 1 mg/kg/day on day 9. On day 21, the patient passed bright red blood per rectum, and her hematocrit dropped to 10%. An angiogram revealed bleeding from the superior mesenteric artery. A resection of the terminal ileum and ascending colon was performed. An ulcerating lesion, 5.5 by 2.5 by 1.5 cm, was found on the mucosal surface at the ileocecal valve. Histopathological examination showed transmural necrotizing granulomatous inflammation and vascular invasion by nonseptate hyphae (Fig. 1D).

View larger version (147K): [in this window] [in a new window]   FIG. 1.   (A) Necrotic lesion on nasal ala prior to surgery. (B) Appearance of patient 1 month after surgery. (C) Branching, nonseptate hyphae of R. schipperae in nasal mucosa. Hematoxylin-eosin stain. Magnification, ×613. (D) Hyphae of R. schipperae in intestinal mucosa. Gomori methenamine-silver stain. Magnification ×613. (E) Thick- and thin-walled chlamydospores of R. schipperae on PDA. Magnification, ×613. Lactophenol cotton blue mount. (F) Cluster of sporangiophores of R. schipperae on CDA. The arrow shows a septate rhizoid. Magnification, ×213. Lactophenol cotton blue mount.

The patient received AMB at 1 mg/kg/day for a total of 40 days and AMB-lipid complex (ABLC) at 5 mg/kg/day intermittently for 12 days during periods of renal insufficiency. Two months after the heatstroke, she has had significant neurological recovery but suffers from dysarthria and ataxia.

	MATERIALS AND METHODS

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Histopathology. Fragments of skin, soft tissue, and cartilage from the right nasal vault and segments of the terminal ileum and ascending colon were submitted for hematoxylin-eosin and Gomori methenamine-silver staining.

Mycology. Samples from the tracheal aspirates, nasal tissue, and small intestine were initially incubated on Sabouraud dextrose agar (SDA) (BBL Microbiology Systems, Cockeysville, Md.) at 25°C. Since the cultures did not sporulate, a small portion of SDA containing fungal growth was excised and incubated in a 10% yeast extract-water culture at 35°C for 10 days in an attempt to promote sporulation (26). Cultures remained sterile, producing only large, thick-walled, globose to somewhat irregularly shaped chlamydospores (Fig. 1E). The nasal isolate was plated on Mycosel agar (BBL), potato dextrose agar (PDA) (Remel, Lenexa, Kans.) (32), and SDA plates for temperature studies at 25, 30, 45, and 50°C. Subcultures of the nasal isolate (UH F 1746) were sent to the Fungal Testing Laboratory, Department of Pathology, University of Texas Health Science Center at San Antonio, for further characterization and designated UTHSC R-3053. There, the isolate was subcultured on Czapek Dox agar (CDA) (Remel) at 30°C, and temperature studies were repeated, but with the addition of 35°C testing.

Antifungal susceptibility testing. The nasal isolate was tested to determine its susceptibilities to AMB, fluconazole (FLU), 5-fluorocytosine (5-FC), and itraconazole (ITRA). Tests were performed by previously described macrodilution methods (22, 33, 43). The minimum lethal concentrations (MLCs) were determined by plating 100-ml samples from the drug-free control tube, the MIC tube, and a tube containing each concentration above the MIC on an SDA plate incubated at 35°C. The MLC was defined as the lowest concentration of antifungal compound resulting in five or fewer colonies on the SDA plate (33).

	RESULTS

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Mycology. Initial cultures on SDA at 25°C produced white, sterile, floccose colonies after 4 days of incubation; the colonies became buff colored at day 6. Temperature studies revealed good growth at 25, 30, 35, and 45°C and no growth at 50°C. The isolate also failed to grow on media containing cycloheximide. Subcultured colonies at 25°C on PDA and SDA were white, floccose in the central portion, and thin and effuse near the periphery. Rapid growth resulting in numerous chlamydospores occurred on both media, but sporangia were lacking. Chlamydospores were thin or thick walled (0.5 to 1 or 3.9 µm wide), globose to subglobose, oval to irregularly shaped, and 10 to 40 µm in diameter (Fig. 1E). A culture in 10% yeast extract-water at 35°C was sterile after 10 days of incubation. Colonies on CDA at 30°C were white, thin, diffuse, and floccose, produced fewer numbers of chlamydospores than at 25°C, but promoted the production of the fruiting structures of R. schipperae Weitzman, McGough, Rinaldi, et Della-Latta (1996). These consisted of up to 10 (usually 4 to 8) sporangiophores arising from a cluster of rhizoids (Fig. 1F, 2, and 3). Mature sporangiophores were brown, being darker at the base, occasionally septate, 150 to 400 µm in length, and 3 to 13 µm in diameter. Sporangia were globose, greyish black, and up to 65 µm in diameter; columellae were subglobose and hyaline (Fig. 4). Sporangiospores were subglobose to oval and faintly striated in water, measuring 4.8 to 6.8 µm long by 3.9 to 5.8 µm wide (Fig. 5). Rhizoids were pale to darker brown, mostly simple (seldom branched), and occasionally septate (Fig. 1F). Some hyphal strands on all media were covered by crystals (Fig. 6). The case isolate was compared with the type isolate (UTHSC 94-538, ATCC 96514, CBS 138.95) obtained from a human lung in San Antonio, Tex., in 1996 (46). On the basis of the above features, the isolate was identified as R. schipperae.

View larger version (124K): [in this window] [in a new window]   FIG. 2.   Cluster of sporangiophores of R. schipperae on CDA. Magnification, ×306. Lactophenol cotton blue mount.

View larger version (96K): [in this window] [in a new window]   FIG. 3.   Rhizoids and sporangiospores of R. schipperae on CDA. Magnification, ×306. Lactophenol cotton blue mount.

View larger version (79K): [in this window] [in a new window]   FIG. 4.   Columella of R. schipperae on CDA. Magnification, ×213. Lactophenol cotton blue mount.

View larger version (114K): [in this window] [in a new window]   FIG. 5.   Columella and striated sporangiospores of R. schipperae on CDA. Magnification, ×613. Lactophenol cotton blue mount.

View larger version (101K): [in this window] [in a new window]   FIG. 6.   Crystals on hyphae of R. schipperae on PDA. Magnification, ×613. Lactophenol cotton blue mount.

Antifungal susceptibility testing. The results of in vitro antifungal susceptibility testing are displayed in Table 1. Based upon drug concentrations normally achievable in patients receiving recommended dosages, the isolate appeared resistant to all antifungal agents tested (AMB, 5-FC, FLU, and ITRA). On the basis of an observed MIC of 4 µg/ml, R. schipperae appears to be somewhat more resistant to AMB than R. arrhizus, the most commonly isolated Rhizopus species, for which AMB MICs are typically 0.125 to 2.0 µg/ml (42). The MICs obtained for the azoles corroborated in vitro data for the type isolate; the MICs of FLU and ITRA were >64 and >8 µg/ml, respectively. The patterns of resistance to azoles observed for this fungus are similar to those seen for other Rhizopus species and zygomycetous genera (25).

	DISCUSSION

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Heatstroke. Heatstroke occurs when exposure to high ambient temperatures produces hyperthermia and neurological dysfunction. Heatstroke produces damage in multiple organs, causing shock, DIC, renal failure, lung injury, lactic acidosis, and electrolyte abnormalities. The mortality rate in heatstroke is 10 to 50%, with 7 to 14% of the survivors suffering permanent neurological impairment (4).

Pathophysiology. The following scenario attempts to explain why this woman experienced heatstroke while her male companions did not and why the uncommon conditions of C. jejuni bacteremia and zygomycosis arose in this patient. The patient's home, San Luis Potosi, lies at an altitude of 1,877 m, and thus, despite its tropical latitude, has a moderate climate (44). Therefore, the patient was not acclimatized prior to departure, increasing the risk of heatstroke (15, 40). Half of her 12-h bus ride was in a non-air-conditioned vehicle; this situation may have also had a detrimental effect on her heat tolerance, because heat stress is cumulative (15).

In vitro antifungal susceptibility. Currently, AMB is the only antifungal agent with clinical utility in the treatment of zygomycosis (25). Although the in vitro data for AMB suggested resistance, the incidence of infections with mold pathogens is sufficiently low that strict correlation of antifungal MICs with clinical results of antifungal treatment has not yet been established (24).

Taxonomy and identifying features. R. schipperae was first described in 1996 after the organism was obtained from bronchial washings and lung specimens from a patient with multiple myeloma in San Antonio, Tex. (46). This case represents only the second isolate of R. schipperae ever described. However, we suspect that the organism may be more common but is not often identified due to the requirement of CDA for sporulation. The organism belongs to the R. microsporus complex, being closely related morphologically to R. microsporus var. microsporus Schipper (1984) Schipper et Sampson (1994) and sharing characteristics with R. caespitosus Schipper et Sampson (1994) in the same complex (35-37) and with Amylomyces rouxii Ellis, Rhodes, et Hasseltine (1976) (9). Characteristics which differentiate these species are displayed in Table 2. Distinguishing features for R. schipperae include its ivory to buff color on PDA, the production of numerous intercalary and terminal chlamydospores on a variety of media, clusters of up to 10 short sporangiophores arising from simple, well-developed rhizoids, the limited range of media for the development of sporangia (especially CDA), and striated sporangiospores. The species is named in honor of M. A. A. Schipper for her work with the Mucorales.

Conclusions. This case exemplifies the severe pathophysiological disturbances that occur in heatstroke with accompanying infectious complications. Heatstroke may alter immune status and predispose to infection by a number of mechanisms: increased translocation of organisms across an ischemic bowel wall, acidosis, complement depletion secondary to DIC, and altered lymphocyte function (5, 8, 11, 13). Heatstroke has previously been recognized as a risk factor for gram-negative bacteremia (11, 17). However, this is the first report of heatstroke as a setting for disseminated zygomycosis. There is one previous example of disseminated infection with the fungus Fusarium oxysporum in a patient with heatstroke (40).