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An aetiopathogenetic analysis of non-endemic nasopharyngeal carcinoma (NPC) in European and Southern American patient groups was performed. Specifically, the study sought to determine the proportion of Epstein-Barr Virus (EBV)-positive tumour cells in NPC patients in two very different populations (Europe and South America) in areas not associated with a high incidence of NPC. Clinical data (age, sex and onset of clinical disease) were also analyzed. A total of 50 NPC samples, 24 from a European hospital (EH) and 26 from two South American hospitals (SAH), were included. Nuclear staining for Epstein-Barr virus-encoded small RNA (EBER) was performed by in situ hybridization (ISH). Latent membrane protein 1 (LMP1) expression was measured by immunohistochemical (IHC) analysis. A higher incidence of NPC was observed in patients > 40 years of age in EH; in SAH, by contrast, the incidence was higher in patients aged ≤ 40 years. Cervical lymph node metastasis was detected in 31 patients (of whom 84.6% were from SAH). A total of 72% of samples were EBERpositive; the incidence of EBER positivity was greater in type 3 NPCs. EBV was detected in a large proportion of epithelial cells in samples from both EH and SAH (75% vs. 69.2%, respectively). An association was found between EBER detection in lymphocytes and patient origin (p = 0.0001). LMP1 expression was detected in 64% of patients. ISH for the detection of EBER is the most sensitive technique for demonstrating EBV in tumour tissue. The incidence of EBV was not significantly greater in either of the study populations, but was significantly higher in patients with type 3 NPC. Definitive histological diagnosis of NPC was reached earlier in EH than in SAH, where metastases were more frequently diagnosed, suggesting that the disease had reached a more advanced stage by the time treatment was started.