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Primary spinal infection (PSI) is a generic term covering a heterogeneous group of infections that can affect the vertebral body, intervertebral disks, the content of the medullary cavity, and adjacent paraspinal tissues. Patients’ characteristics can vary significantly, notably according to their age, and some of these characteristics undoubtedly play a primordial role in the occurrence of a PSI and in the type of offending pathogen. Before approaching the subject of laboratory diagnostics, it is essential to define the characteristics of the patient and their infection, which can then guide the physician toward specific diagnostic approaches. This review critically examined the roles and usefulness of traditional and modern laboratory diagnostics in supporting clinicians’ decision-making in cases of pediatric and adult primary spinal infection (PSI). It appears impossible to compare PSIs in children and adults, whether from an epidemiological, clinical, bacteriological, or biological perspective. The recipients are really too different, and the responsible germs are closely correlated to their age. Secondly, the interpretation of traditional laboratory blood tests appears to contribute little guidance for clinicians attempting to diagnose a PSI. Biopsy or needle aspiration for bacterial identification remains a controversial subject, as the success rates of these procedures for identifying causative organisms are relatively uncertain in pediatric populations.Using nucleic acid amplification assays (NAAAs) on biopsy samples has been demonstrated to be more sensitive than conventional cultures for diagnosing PSI. Recent advances in next-generation sequencing (NGS) are particularly interesting for establishing a microbiological diagnosis of a PSI when standard cultures and NAAAs have failed to detect the culprit. We can even imagine that plasma metagenomic NGS using plasma (known as “liquid biopsy”) is a diagnostic approach that can detect not only pathogens circulating in the bloodstream but also those causing focal infections, and thus eliminate the need for source sample collection using costly invasive surgical procedures.