Abstract Identifying differences in a key social trait between two populations of die same species is important for understanding the evolution of sociality. Previous studies of new colonies in Exoneura bicolor, an Australian allodapine bee, have shown that there are high levels of kin cofounding in a montane population. The only study to examine intra-colony relatedness in a heathland population has found that new multifemale colonies are not formed by kin. In this study we used an experiment to investigate both cofounding behaviour and intra-colony relatedness in E. bicolor from a heathland population. Nest substrate was placed either 0.05 or 1 m distant from source nests in a novel environment. Although there were no differences in cofounding rate or intra-colony relatedness between die two treatments there was, overall, a high rate of cofounding: 53% of new nests were multifemale, approximately twice as high as found in previous field-based studies. Relatedness among cofoundresses was not only different from zero, r± SE = 0.597 ± 0.097, but almost identical to that found in montane populations. A constraint, such as nest substrate distribution, is suggested as a proximate factor affecting the expression of cofounding behaviour in E. bicolor. The implication of such proximate constraints for inferring the phylogenetic origins of social behaviour is discussed.