A 49-year-old female with a past medical history of severe asthma and osteoporosis presented to the Department of Allergy and Clinical Immunology with a 2-week history of arthralgia of the left hip and knee and a 6-day subjective fever, with a maximum body temperature of 39.6 °C. The patient had been on high-dose inhaled corticosteroids (ICS) fluticasone propionate and long-acting β2-agonist (LABA) formoterol fumarate (500 μg of fluticasone and 50 μg of formoterol twice daily) and low-dose (6 mg daily) oral corticosteroids (OCS) for more than 10 years, and she had previously undergone surgical treatment for lumbar osteoporosis combined with pathological fracture. Additionally, the patient denied any family medical history resembling her symptoms or other inherited disorders. At admission, the patient presented with a full moon face, buffalo hump, centripetal obesity, and stable vital signs. The physical examinations were grossly normal, except for percussion pain in the left hip. Routine laboratory tests were significant for elevated blood leukocyte count at 25.70 × 109/L (4.0–10.0 × 109/L), neutrophil count at 21.7 × 109/L (1.8–8.0 × 109/L), neutrophil ratio at 84.3% (40.0–70.0%), procalcitonin (PCT) at 0.19 ng/mL (0–0.05 ng/mL), and erythrocyte sedimentation rate (ESR) at 90 mm/h (0–20 mm/h). Peripheral T-lymphocyte subpopulation and absolute count manifested decreased levels of suppressor T-lymphocyte (TS), B-lymphocyte, and natural killer (NK) cells. Blood and sputum cultures for bacterial pathogens were all negative, as were serologies for common respiratory pathogens. X-rays revealed degeneration and osteoporosis of the left hip and the left ankle, while chest CT did not show any significant abnormalities.

The initial consideration was given to infection because of the patient's febrile symptoms and elevated inflammatory markers. Peripheral blood culture was obtained, and simultaneously, the patient was started on intravenous (i.v.) piperacillin-Sulbactam (3.0 g, Q8h) as the empirical anti-infective therapy. After 6 days of treatment, the patient was still feverish (temperature range between 38 and 39 ℃) and experiencing joint pain (mainly in the left hip, knee, and ankle), which may temporarily resolve after the fever had subsided. A multidisciplinary consultation was then convened, and the diagnostic possibilities of steroid-related osteoporosis, osteomyelitis, and infectious fever were discussed. In the meantime, brucellosis infection could not be excluded as the patient's clinical features were similar to the typical symptoms of brucellosis infection: undulant fevers, sweating, and migratory arthralgia and myalgia [10, 11]. While awaiting blood culture results, the anti-infective regimens were subsequently adjusted to i.v. meropenem (1 g, Q8h) and oral minocycline (100 mg, BID). After 5 days of adjusted treatment, the patient remained febrile, but her daily fever spikes showed a decreasing tendency (daily peak body temperature from 39.1℃ to 37.5℃). However, during this phase, the root cause of her fever was still not well known. Multiple blood and sputum bacterial cultures were negative, and magnetic resonance imaging of lower extremity joints revealed no evidence of osteomyelitis.

In order to identify the pathogen, conventional diagnostic tests and metagenomic next-generation sequencing (m-NGS) were performed simultaneously. m-NGS was performed with blood drawn at the peak of the febrile phase and detected C. koseri with 979 reads and a relative abundance of 38.36% (Table 1). Afterward, whole-body 18F-fluorodeoxyglucose positron emission tomography/computed tomography (18F-FDG PET/CT) was conducted to identify the infection foci. The PET/CT scan manifested a tissue-like mass with significantly increased FDG uptake in the left middle and lower abdomen. The delayed scan indicated a further increase in the lesion metabolism, suggesting a high possibility of infection while not ruling out the possibility of tumor (Fig. 1). The patient then underwent ultrasound-guided needle aspiration biopsy to define the nature of the FDG-avid lesion and to rule out the possibility of malignancy. The abdominal biopsy tissue was used for m-NGS, pathogen culture, and mass spectrometry (MS; Biomerieux Vitek, France). The m-NGS results of the biopsy tissue from the abdominal mass indicated C. koseri infection, with 263, 387 valid sequence reads and a coverage of 88.1636% (Fig. 2A, Table 1). This result echoes the m-NGS detection of blood sample. For pathogen culture, the colonies displayed a consistent morphology after inoculation, which was subsequently identified by mass spectrometry and also turned out to be C. koseri (Fig. 2B). Pathological analysis did not reveal tumor cells. In summary, C. koseri infection was confirmed by the mutual validation of blood m-NGS, biopsy tissue m-NGS, biopsy tissue culture, and mass spectrometry results. The antimicrobial susceptibility results indicated that the bacteria were susceptible to meropenem, ertapenem, ceftriaxone, amoxicillin-clavulanate, and levofloxacin. To explore the source of the infection, the patient subsequently received a colonoscopy, which did not demonstrate any structural abnormalities. After consulting the surgical department, conservative management was considered. Following confirmation of C. koseri infection, i.v. meropenem (1 g, Q8H) was continued to use as targeted antimicrobial therapy, while minocycline was discontinued. After receiving 12 days of continuous medication, the patient's fever subsided, and her arthralgia was also alleviated. The patient was discharged from the hospital and continued to be treated orally with faropenem (0.3 g, Q8h) for 2 months. During the 8-month follow-up, abdominal CT showed gradual lesion absorption (Fig. 3).

The coronal and sagittal section on abdominal 18F-FDG PET/CT. 18F-FDG PET, CT on the soft-tissue window, and fused PET/CT images demonstrated FDG accumulation in the left middle and lower abdomen. The hypermetabolic areas (yellow-white) indicated that the FDG-avid soft tissue mass was in an active inflammatory process

m-NGS A and mass spectrometry B analyses of abdominal biopsy tissue indicated C. koseri infection. A: The C. koseri genome sequences mapping by m-NGS. The result of mNGS showed 263387 reads corresponding to the C. koseri, with a coverage of 88.1636%. The X-axis represents the genome size of C. koseri and the Y-axis represents the number of sequences detected within different genomic segments. B: Mass spectrum of C. koseri. MALDI-TOF mass spectrometry showed the specific spectrum, which matched the registered pattern of C. koseri. The X-axis represents the mass-to-charge ratio (m/z) of different peptide fragment ions and the Y-axis represents the ion intensity. Peaks at specific m/z values indicate the presence of corresponding ions in the sample. The intensity of each peak reflects the abundance of the corresponding ion. m-NGS metagenomic next-generation sequencing, MALDI-TOF mass spectrometry matrix-assisted laser desorption ionization time-of-flight mass spectrometry

Comparison of the abdominal CT results before and after treatment. Serial CT scan images showed progressive resolution of the lesion in the left middle and lower abdomen over the course of 2 months on antimicrobial therapy: A prior to treatment; B 1 month after discharge; C 4 months after discharge; D 8 months after discharge