Attachment is an organizational construct that serves to organize the development of emotional bond between human infants and their caregivers (Bowlby, 1958). In early infancy its function is to obtain protection and care from another person by adapting one's behavior to characteristics of the key attachment figure (Bowlby, 1969). This early development results in different attachment styles that can be assessed in terms of two dimensions of security/insecurity: attachment-related anxiety and attachment-related avoidance (e.g., Ainsworth et al., 1978; Brennan et al., 1998; Fraley and Spieker, 2003). Attachment styles have well-documented cognitive, physiological, and neurological correlates (e.g., Diamond, 2001; Gillath et al., 2005), and behavioral and psychological consequences that last into adulthood, including self-regulation of stress and emotions, influence on relationship quality with romantic partners, sexual motivation, and reactions to relationship breakups or losses (see Shaver and Clark, 1994; Mikulincer et al., 2003; for reviews).

Most studies have focused on the environmental effects shaping attachment (such as parental behavior, Fearon et al., 2014). more recently however, candidate gene studies have reported associations between attachment styles of human infants and polymorphisms in their dopamine D4 receptor, serotonin transporter, and oxytocin receptor (OXTR) genes (Lakatos et al., 2000; Barry et al., 2008; Chen et al., 2011; Spangler, 2011), suggesting that genetic polymorphisms may moderate the links between parental behavior and other environmental effects and infant attachment. Therefore, it has become obvious that attachment styles are shaped by a combination of genetic factors and social experiences (Fonagy, 2001). This is well-demonstrated by the fact that carrying a specific genetic polymorphism can be associated with developing a particular attachment style in one kind of social environment but not in another (Gillath et al., 2008). For instance, Chen et al. (2011) found that the A allele, as compared to the G allele, of OXTR rs2254298 was more likely associated with secure attachment in a non-Caucasian sample but not in a Caucasian sample. Such results in human infants allow for limited conclusions however, because of the genetic relatedness of the infants and their parents. Allelic variations associated with different attachment styles of infants have been shown to affect also various characteristics of the parents, allowing for an alternative, indirect, link between genotype and infant attachment.

In human subjects variations of oxytocin-related genes have been found in association with various personality traits, their components and neurological correlates, such as agreeableness extraversion, social loneliness, anxiety, and amygdala volume (Lucht et al., 2009; Saphire-Bernstein et al., 2011; Haram et al., 2014; Wang et al., 2014). Nonhuman examples also can be found, namely cats with the A allele in the SNP G738A show significantly higher “Roughness” personality scores than cats without the A allele (Arahori et al., 2016). Parental personality affects parenting style (Council et al., 1988; Kendler et al., 1997; Metsäpelto and Pulkkinen, 2003), thereby personality traits have an impact on the quality of parent-child relationships, and on the children's attachment style (Kochanska et al., 2004). However, as infants and their parents likely carry similar alleles in the polymorphic regions of their OXTR, in humans it is difficult to dissect whether and how infant genotypes, parent genotypes and other characteristics of parents (e.g., their personality or their own attachment style) affect infant attachment. The domestic dog, however, provides a unique opportunity to investigate this question.

Dogs have been part of human societies for longer than any other domestic species (Clutton-Brock, 1999). Their ability to form attachment with humans is one of the most widely recognized consequences of domestication (Topál and Gácsi, 2012). Topál et al. (1998) were the first to reveal that dogs develop attachment to their owners analogous to the infant-mother attachment in humans (for a replication see Prato-Previde et al., 2003).

Similarly to human infants, the most widely used paradigm to investigate dogs' attachment behavior toward their owners is the Strange Situation Test (SST; Topál et al., 1998). The test consists of seven episodes, each lasting 2–3 min, when the dog is either with the primary caregiver (owner), with a stranger, or alone in an unfamiliar place. The essential element of the test is that separation from the human caregiver in an unfamiliar environment evokes moderate stress, which manifests in proximity seeking while the reunion with the caregiver evokes contact-seeking behaviors. Multivariate analysis of Topál et al. data (1998) (factor and cluster analyses) separated three key aspects of dogs' behavioral structure. These major factors revealed that dogs' behavior during the test was affected by: (i) their sensitivity to the separation from the owner (Attachment), (ii) the degree of stress the unfamiliar environment evoked from them (Anxiety), and (iii) their responsiveness to the stranger (Acceptance). Their study demonstrated that adult dogs show specific patterns of attachment behavior toward their owners, and dogs' individual behavior patterns can be explained by the different combinations of these determining factors. Many studies have reported that dogs show a great variability in their behavior in the SST (Topál et al., 1998; Gácsi et al., 2001; Naderi et al., 2002; Prato-Previde et al., 2003; Mariti et al., 2014; Scandurra et al., 2016) suggesting that adult dogs are particularly suitable subjects for studying the phenomenon of animal-to-human attachment.

Pet dogs also offer a good model for investigating to what extent attachment patterns are shaped by the independent genetics of the dogs and their owners and by environmental factors, such as the owners' personality, attachment style, or the country they live in. Drawing a parallel between infant-mother and dog-owner attachment has recently gained further support from the finding that ownership styles seem to be composed of components similar to those of human parenting behavior, suggesting that owners largely use their parenting repertoire when interacting with their dogs (Cimarelli et al., 2017). Importantly, however, only little (and indirect) data is available on how different genetic and environmental factors influence dogs' attachment to human.

Similarly to the finding that polymorphism in the OXTR gene is related to security/insecurity of mother-infant attachment in humans (Chen et al., 2011), it has been suggested that oxytocin plays an important role in the relationships between dogs and their owners, with higher oxytocin levels being associated with a more positive relationship from perspective of the owner (Thielke and Udell, 2015). Thielke and Udell even suggested that intranasal OXT administration can be combined with behavioral therapies for dogs with behavioral problems related to separation anxiety. Somewhat contrasting this view, van Rooy et al. (2015) in a study examining several candidate genes (OPRM1, AVPR1A, DRD2, OXTR) and their associations with separation-related distress in 42 Australian golden retrievers, found no significant associations between separation-related behavior scores and OXTR gene.

The main purpose of the present study was to explore environmental and genetic influences on dogs' attachment behavior. In order to benefit from the genetic unrelatedness of dogs and their owners, in separate analyses we investigated (1) whether various OXTR polymorphisms of dogs as well as their owners are associated with the attachment behavior of the dogs in two different countries, (2) whether such effects of the dogs' genotypes are affected by the age, sex, and neutering of the dogs. Finally, to tackle potential mechanisms that may mediate the effects of owner genotypes on dog attachment, we analyzed (3) if owner personality, attachment style, and attachment to pets have an effect on dogs' attachment behavior.

Chen et al. (2011) had suggested that one source of the variation in human infants' attachment to their mothers is the polymorphism of their OXTR gene but they remained cautious about this conclusion due to the genetic relatedness of infants and their parents. Based on the analogy between infant-mother and dog-owner attachment, our findings seem to confirm their suggestion, as the present study provides the first evidence that genetic variations in dogs' OXTR gene are associated with their attachment behavior to their owners. All behavioral aspects measured in the SST (Attachment, Anxiety, and Acceptance) showed significant association with all three dog OXTR SNPs investigated in this study (as a main or an interaction effect). All 3 canine OXTR SNPs have proved to have behavioral associations also in former studies. Behavioral associations of the −213AG polymorphism have already been reported in other studies. The −213AG polymorphism had been shown to be associated with proximity seeking not only in Border Collies but also in German Shepherds (Kis et al., 2014). Another study (Kovács et al., 2016b) investigating dogs' social sensitivity has also found significant main effect of the same OXTR polymorphism (−213AG) on readiness to look at the human face in two test situations. Moreover, Turcsán (2017) found that this SNP was associated with greeting behavior, especially when the dog had no prior negative experience with the experimenter. Oláh et al. (2017) also found that dogs' first reaction and their friendliness in the threatening approach test were significantly modulated by −213AG and −74GC polymorphisms. In line with our finding that −213AG polymorphism is associated with Stranger acceptance, Romero et al. (2014) also found that oxytocin modulates social motivation to approach and affiliate with conspecifics and human partners. Earlier studies (Windle et al., 1997; Neumann, 2002; Bello et al., 2008) had also shown, that oxytocin can regulate the activity of the hypothalamic–pituitary–adrenal axis, thereby modifying the stress response. This mechanism may explain the associations we found between the OXTR polymorphisms and Anxiety composite score.

We found some differences between the results of different models. For example −74CG has significant main effects on dogs' Attachment, Anxiety as well as Acceptance scores in our first model (when testing its effects in interaction with the owners' genotypes), but in the second model it has an influence only on dogs' Anxiety and only when tested in interaction with dogs' sex. Moreover, while −74CG (and not −213AG) has a main effect on dogs' Anxiety in the first model,−213AG (and not −74CG) has a main effect on the Anxiety in our second model. This seems to result from the facts that sex of the dogs influenced the associations of two canine OXTR SNPs, namely the effects of −213AG and −74CG on dogs' Anxiety. It has been shown both in humans (e.g., Herzmann et al., 2013) and dogs (Nagasawa et al., 2015; Kovács et al., 2016a; Turcsán, 2017) that oxytocin can have differential effects on males and females. In prairie voles more is known about the underlying mechanisms of such sex differences, as Smeltzer et al. (2006) found higher binding by the oxytocin receptors in the medial prefrontal cortex in female than in male prairie voles. Another possible explanation is that steroid hormones, such as estradiol and progesterone, can modulate the affinity of the oxytocin receptors (estradiol enhances OXT receptor affinity, while progesterone has been shown to decrease receptor binding; Gimpl et al., 2002; Choleris et al., 2008).

Another important finding of the study is that both dog and owner OXT genetic variation shapes the dog-owner attachment in an interactive manner. Earlier research has also shown a mutual effect of both dogs and their owners on the peripheral oxytocin levels of both parties (Nagasawa et al., 2009, 2015). This is the first study, however, providing evidence that the oxytocin system of both parties impacts on dogs' attachment behavior. We found significant effects of two human OXTR SNPs (rs2254298 and rs1042778) on the Attachment composite score and one SNP (rs53576) on Acceptance, and several significant interactions of the effects of the human and dog OXTR gene on the attachment behavior of dogs (e.g., −213AG × rs2254298,−74CG × rs53576,−94TC × rs1042778,−74CG × rs1042778). Dog owners' behavior is likely to be one of the greatest environmental factors influencing the dogs' attachment behavior. It is possible that the owners' behavior mediates this link between the owners' genotype and the dogs' relationship to them, particularly given Bakermans-Kranenburg's and van Ijzendoorn (2008) finding that OXTR is related to parenting style in humans. As dogs are unrelated to their human caregivers, the owner's genetic background may have an influence on their parenting style or other relevant behavior that, in turn, through epigenetic processes, affects the dogs' attachment behavior or the effects of the dogs' own OXTR genotype on it. However, given that in this study a considerable number of human DNA samples were missing, future studies with larger sample sizes need to consolidate these results.

As a further confirmation of owner influences on dog attachment, we have also found that the owners' Bond-related avoidance to their partners and Pet-related avoidance influenced all the three composite scores of their dogs' attachment, Openness and Pet-related anxiety affected dogs' Anxiety and Acceptance, while Extraversion personality trait influenced the Anxiety composite score. Previously, Konok et al. (2015) examined in a questionnaire study whether owners' attachment style and personality traits influence the occurrence of separation-relation disorder in the dogs. They found that owners scoring higher on self-reported attachment avoidance are more likely to have dogs with separation-related disorder. They suggested that owners' attachment style influences their caregiving behavior toward their dogs, and owners with attachment avoidance may show less consistent responsiveness to their dog's needs. Schöberl et al. (2016) also investigated the effects of owner personality and found that owners with high neuroticism and agreeableness had dogs with lower cortisol reactivity in the Strange Situation Test. In contrast, in humans, children's insecure attachment, behavior problems and separation anxiety disorders are often associated with mothers' neuroticism and anxiety disorder (Manassis et al., 1994; Biederman et al., 2001; Kochanska et al., 2004). Interestingly, in our study the Neuroticism personality trait associated with none of the dog composite scores. Our finding about the correlation between owners' personality and their dogs' behavior is especially important from an applied perspective, as environmental factors may have the potential to modify oxytocin-related behavioral changes.

Finally, one of the most powerful effects we found was a difference between Austria and Hungary. There were main effects of country on two of the three behavioral components (Attachment and Anxiety composite scores) with dogs in Hungary showing higher Attachment and Anxiety, than dogs in Austria. Country has also influenced the effect of dog genetic background on attachment; similarly to Chen's et al. (2011) study where a certain OXTR genotype was associated with secure attachment in a non-Caucasian sample but not in a Caucasian sample. Oxytocin functions, similarly to serotonin (Yoshida et al., 2009) and dopamine functions (Liu and Wang, 2003), are likely be influenced by multiple factors, including other genetic polymorphisms that vary across ethnic populations (e.g., Chang et al., 1996; Kunugi et al., 1997). These uncontrolled genetic differences in our study could potentially mask the function of oxytocin and thus may have weakened the measurable association between variations in OXTR function and attachment behavior of dogs. Alternatively or additionally, different components of the dogs' social environment may differ between the two countries. It has been suggested that the behavioral expression of certain genotypes is sensitive to input from the social environment (Way and Taylor, 2010). Kim et al. (2010) suggested that the social environment can alter or even reverse the phenotypic expression of different genotypes. They found that culture-specific norms as a form of social input can also affect phenotypic expression of OXTR.

In conclusion, our study provides experimental evidence that genetic variations of the OXTR gene in both dogs and their owners, as well as various aspects of dogs' environmental background are associated with their attachment to their human caregivers. Based on previous and present results we propose that polymorphism in the oxytocin receptor gene is a potentially important factor in regulating dog-human relationship. Although the complex joint effects of genetic and environmental factors on dogs' human-directed social behavior warrant further investigation, these findings offer a promising approach to studying causes and treatments of separation anxiety in dogs.

KK, ZV, AK, and JT: Designed the study. All authors prepared the study material and data acquisition. KK and DK: Entered the data and prepared it for statistical analyses; KK, ZV, AK, BT, and JT: Analyzed the data; KK, ZV, and JT: Interpreted the data; ZV, ZR, and JT: Obtained funding; KK: Wrote the first draft of the manuscript; KK, ZV, AK, MG, and JT: Critically revised the manuscript for important intellectual content. All authors gave final approval of the manuscript version to be published and agreed to be accountable for all aspects of the work in ensuring that questions related to the accuracy or integrity of any part of the work are appropriately investigated and resolved.