Identification of Mycoplasma hominis Infection by Nanopore 16S rDNA Amplicon Sequencing in Epidural Empyema After Neurovascular Surgery

Dear Editor,

Mycoplasma hominis is a common genitourinary and respiratory tract colonizer that is occasionally deadly in immunocompromised and neonatal patients.1, 2 As one of the smallest bacteria lacking a cell wall, M. hominis is challenging to identify using Gram staining or conventional culturing.3, 4 This means that extragenitourinary M. hominis infection of healthy individuals can easily be misdiagnosed. High-throughput nanopore 16S rDNA amplicon sequencing has recently enabled pathogens to be detected with high sensitivity and specificity.5 We present a case of a neurosurgical wound infected with M. hominis that was diagnosed using nanopore 16S rDNA amplicon sequencing.

A 52-year-old male visited our institution for two incidentally found unruptured intracranial aneurysms of the left middle cerebral artery (MCA). Although asymptomatic, a surgical intervention was decided since they were large (7 mm and 9 mm) and of increasing size. The patient underwent superficial temporal artery-MCA double-barrel bypass surgery. An anastomosis was successfully formed but brain swelling was noticed after declamping, and so the operation site was closed without replacing the bone flap.

The patient became febrile (38.2℃) immediately after the surgery, followed by marked elevation of C-reactive protein (CRP) (22.79 mg/dL) on postoperative day (POD) 2, suggesting a postoperative infection (Fig. 1). Empirical pip/tazo (piperacillin/tazobactam) targeting microaspiration pneumonia was started on POD 3 but discontinued on POD 5 after negative results in aspirate culturing and the absence of relevant clinical features. Notably, urinalysis revealed the presence of hematuria, with a red blood cell count (RBC) of 1,124/µL. However, the white blood cell count in the urine was within the normal range of 1–4/µL, and no bacteria were detected. Both urine and blood cultures yielded negative results. The surgical wound appeared clean, and cerebrospinal fluid cultures as well as 16S rDNA polymerase chain reaction (PCR) did not reveal any signs of infection.

Fig. 1
Timeline of the clinical course and diagnostic test results. A: Relationship between the change in infection indices (WBC, CRP, and body temperature) and the use of antibiotics during hospitalization. B: Gadolinium-contrast-enhanced MRI, with epidural empyema indicated by an arrow. C: Results of nanopore 16S rDNA amplicon sequencing. CRP, C-reactive protein; MRI, magnetic resonance imaging, Op., operation; Pip/tazo, piperacillin/tazobactam; POD, postoperative day; WBC, white blood cell count.

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The spiking fever of 38℃ returned on POD 6, accompanying leukocytosis and CRP elevation. A small amount of oozing was noticed at the wound, which soon turned into a pus-like discharge. Brain magnetic resonance imaging showed extensive epidural empyema at the surgical site, for which wound revision was decided on POD 10. A moderate amount of purulent fluid was observed within the epidural space and subcutaneous layer. Pus was collected for testing, followed by massive irrigation.

Despite strong suspicions of infection, all conventional diagnostic methods yielded negative results, including Gram staining, AFB tests, and pus culturing. Meanwhile, 16S PCR revealed a strong positive band on electrophoresis, and so nanopore sequencing was performed from the PCR product as described previously.6, 7 After 16S sequencing, M. hominis was identified as the pathogen within 1 day, and the sequenced reads did not align meaningfully with other known bacteria, ruling out the possibility of a polymicrobial infection. Consequently, treatment with levofloxacin was initiated on POD 13, which led to the resolution of fever and headache within 2 weeks. The patient was discharged without any neurological complications.

The application of 16S sequencing in this case enabled definitive and rapid pathogen identification of M. hominis. This sequencing technique has demonstrated its utility in identifying pathogens following neurosurgical procedures, particularly in cases where infections become culture-negative after initiating antibiotic therapy.8, 9 Wound infections caused by M. hominis are rare, and notorious for their low positivity rates in conventional cultures.1, 4 Central nervous system infections caused by M. hominis have been documented in only 21 adult cases between 1950 to 2022, but the actual incidence remains unknown.8, 9 The exact route of dissemination to the wound is yet to be elucidated, although the evidence is strongest for the commensal bacteria spreading hematogenously through natural barriers disrupted by procedures such as urinary catheterization, endotracheal intubation, or suction.8 Our patient underwent both Foley catheter insertion and endotracheal intubation as preoperative procedures. The high RBC in the urine strongly suggested that traumatic catheter insertion had introduced the commensal M. hominis into the bloodstream, ultimately leading to its dissemination and subsequent infection at the neurosurgical site. Performing 16S sequencing on urine or blood samples might have provided further insight into this hypothesis, but we did not have access to the necessary samples to conduct additional testing.

In conclusion, the findings in this case highlight the importance of considering M. hominis as a rare potential cause of neurosurgical wound infections, particularly following procedures involving the manipulation of mucosal barriers. The application of 16S amplicon sequencing proved highly beneficial in detecting uncommon bacteria that are typically missed in conventional culturing. Further research is warranted to better understand and delineate the exact role and route of dissemination of M. hominis in neurosurgical wound infections.