Studies using teeth to estimate age in marine mammals presume that the neonatal line (NNL) develops at birth. This study of Indo-Pacific bottlenose dolphins (Tursiops aduncus (Ehrenberg, 1833)) is the first to investigate when the NNL appears in odontocete dentine. Two to four teeth were prepared by decalcification, thin-sectioning, and staining for 103 dolphins, including 7 dolphins of known age. Tooth length, prenatal and postnatal dentine and NNL widths were measured. Developmental class (foetus, young neonate, older neonate, <1-year-old calf, 1-year-old calf) was assigned using carcass external features. NNL presence or absence was categorised for individual dolphins. The NNL was absent in a near-term foetus and all except one young neonate and fully formed in 50% of older neonates, whose estimated ages were 1 week to 2 months. It was absent in a known-age dolphin aged 4–7 weeks. NNL width was greater in dolphins less than 1 year old compared with those that were 1 year old. Factors that trigger NNL development are unknown. The present study suggests that the NNL may not be related to birth per se in dolphins, as has been clearly demonstrated in humans. Physiological processes, driven by diet, and behavioural changes during the first few months of postnatal life may be important drivers for NNL formation in odontocetes.

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