3055 INTRODUCTION Although nestmate recognition in social insects has been widely studied, previous work has used a variety of methodologies and social contexts. Bioassays to quantify the differential treatment of nestmates versus non-nestmates range from presenting natural guards at colony entrances with incoming foragers to observing isolated non-guard individuals in containers and presenting them with individuals who have been isolated from a colony since eclosion. The use of these highly artificial contexts has the advantage of affording greater control of the experimental conditions, and such studies have generated significant advances in our understanding of group-level behavioural discrimination (Breed, 1983; Getz and Smith, 1983; Breed et al., 1988; Buchwald and Breed, 2005). However, this may be at the expense of relevance to the actual challenges faced by discriminators in nestmate recognition. The theory of optimal acceptance thresholds predicts that a discriminator will behave more or less permissively depending upon the context, by trading off one error (reject nestmate) for the other (accept non-nestmate) to minimize the total cost of all errors (Reeve, 1989; Couvillon et al., 2009; Jones et al., 2012). One such context that affects recognition behaviour is the perceived threat: for example, when honey bees face increased levels of conspecific robbing, the guards’ threshold becomes less permissive, resulting in a lower proportion of both non-nestmates and nestmates being accepted (Downs and Ratnieks, 2000; Couvillon et al., 2008). Another context is perceived proximity of the colony; paper wasps’ aggression varies depending on the presence of a nestmate or a familiar nest fragment (Starks et al., 1998). However, in spite of these data, many previous studies that quantify recognition behaviour do not account for context. Additionally, results obtained in different contexts are often compared across studies (Johnson et al., 2011). If context is not considered, such comparisons are rendered meaningless. In this study we carried out bioassays on nestmate recognition by guards in two species of eusocial bees, the honey bee Apis mellifera Linnaeus 1758 and the stingless bee Tetragonisca angustula (Latreille 1825). We systematically varied the naturalness of the context from fully natural (guards at nest entrances) through different degrees of unnatural (single versus two guards in a container, with and without colony odour). If there is no effect of context on guarding recognition behaviour (accept/reject an introduced bee), we would expect guards to make similar recognition errors under all treatments. Alternatively, if context does play a role, then we would expect guards to make different recognition errors between contexts. Specifically, we predicted that the lowest total recognition errors would be at the natural nest entrance, with the other contexts varying in their degree of error. SUMMARY Nestmate recognition studies, where a discriminator first recognises and then behaviourally discriminates (accepts/rejects) another individual, have used a variety of methodologies and contexts. This is potentially problematic because recognition errors in discrimination behaviour are predicted to be context-dependent. Here we compare the recognition decisions (accept/reject) of discriminators in two eusocial bees, Apis mellifera and Tetragonisca angustula, under different contexts. These contexts include natural guards at the hive entrance (control); natural guards held in plastic test arenas away from the hive entrance that vary either in the presence or absence of colony odour or the presence or absence of an additional nestmate discriminator; and, for the honey bee, the inside of the nest. For both honey bee and stingless bee guards, total recognition errors of behavioural discrimination made by guards (% nestmates rejected + % non-nestmates accepted) are much lower at the colony entrance (honey bee: 30.9%; stingless bee: 33.3%) than in the test arenas (honey bee: 60–86%; stingless bee: 61–81%; P<0.001 for both). Within the test arenas, the presence of colony odour specifically reduced the total recognition errors in honey bees, although this reduction still fell short of bringing error levels down to what was found at the colony entrance. Lastly, in honey bees, the data show that the in-nest collective behavioural discrimination by ca. 30 workers that contact an intruder is insufficient to achieve error-free recognition and is not as effective as the discrimination by guards at the entrance. Overall, these data demonstrate that context is a significant factor in a discriminators’ ability to make appropriate recognition decisions, and should be considered when designing recognition study methodologies. Key words: Apis mellifera, Tetragonisca angustula, acceptance threshold, eusocial bee, social insects. Received 13 January 2013; Accepted 14 April 2013 The Journal of Experimental Biology 216, 3055-3061 © 2013. Published by The Company of Biologists Ltd doi:10.1242/jeb.085324 RESEARCH ARTICLE Context affects nestmate recognition errors in honey bees and stingless bees Margaret J. Couvillon 1, *, Francisca H. I. D. Segers 1,2,3 , Roseanne Cooper-Bowman 1 , Gemma Truslove 1 , Daniela L. Nascimento 2 , Fabio S. Nascimento 2 and Francis L. W. Ratnieks 1 1 Laboratory of Apiculture and Social Insects, School of Life Sciences, University of Sussex, Falmer, Brighton, BN1 9QG, UK, 2 Departamento de Biologia, Faculdade de Filosofia, Ciências e Letras de Ribeirão Preto, Universidade de São Paulo, CEP 14040-901, Ribeirão Preto, SP, Brazil and 3 Laboratoire d’Ethologie Expérimentale et Comparée, EA4443, Université Paris 13, Sorbonne Paris Cité, 99 Avenue J. B. Clément, 93430 Villetaneuse, France *Author for correspondence (M.Couvillon@Sussex.ac.uk) THE฀JOURNAL฀OF฀EXPERIMENTAL฀BIOLOGY