In children with suspected mitochondrial disease, including those being evaluated for conditions such as thymidine kinase 2 deficiency (TK2d), muscle biopsy and structured diagnostic criteria remain important tools, especially when genetic testing is inconclusive, according to results published recently in Neuromuscular Disorders.
A large retrospective study found that the modified Walker criteria were more sensitive than the Nijmegen criteria for identifying primary mitochondrial disease, while both maintained high specificity. These results reinforce that traditional approaches still matter in the era of advanced sequencing. The Walker and Nijmegan criteria are specialized, evidence-based diagnostic systems for identifying primary mitochondrial diseases, especially in children.
Researchers at Oulu University Hospital reviewed 220 pediatric patients who underwent muscle biopsy for unexplained neuromuscular symptoms between 1990 and 2024. A confirmed genetic diagnosis was made in 58 patients (26%).
Of these, 12 (21%) had primary mitochondrial diseases, 17 (29%) had genetically defined diseases with secondary mitochondrial dysfunction and 29 (50%) had other neuromuscular disorders. Despite advances in testing, 162 of 220 patients (74%) remained without a confirmed genetic diagnosis at the time of analysis, although the proportion of undiagnosed cases declined over time.
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Using a “definite” cutoff, the modified Walker criteria showed 75% sensitivity and 100% specificity among patients with known genetic causes. In comparison, the Nijmegen criteria had 50% sensitivity and 98% specificity.
Positive predictive values were high at 100% for modified Walker and 86% for Nijmegen. When probable and definite scores were combined, sensitivity reached 100% for modified Walker and 75% for Nijmegen. Patients with primary mitochondrial disease had significantly higher Nijmegen scores than all other groups.
Muscle biopsy findings also provided meaningful information. Abnormalities were seen in 34% of patients on light microscopy and mitochondrial changes were detected in 28% on electron microscopy. Decreased oxidative phosphorylation enzyme activity was identified in 49 of 189 tested patients (26%), although only 13 of those had a confirmed genetic diagnosis.
“These results underscore the continued relevance of muscle biopsy and functional studies on patient-derived samples in selected cases, particularly when variants of uncertain significance are identified,” explained this study’s authors.
Importantly, secondary mitochondrial dysfunction was frequent in non-mitochondrial genetic diseases, highlighting the complexity of interpreting abnormal results in disorders that may resemble TK2d or other primary mitochondrial conditions.
For patients and families, these results mean that a normal or unclear genetic test does not end the diagnostic journey. Muscle biopsy and clinical scoring systems can still help clarify whether a child’s symptoms are due to a primary mitochondrial disease such as TK2d or another condition with secondary mitochondrial involvement. This layered approach may guide treatment decisions, inform prognosis and support more accurate counseling while research continues to identify new genetic causes.
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