1 Delimitation of Call Types of Red Crossbill (Loxia curvirostra) in the 2 Western Palearctic 3 Ralph Martin1, Julien Rochefort2, Roger Mundry3, Gernot Segelbacher1 4 5 1 University of Freiburg, Tennenbacher Strasse 4, 79106 Freiburg, Germany; 2 12 rue de 6 la Tourelle, 91600 Savigny-sur-orge, France; 3 Max Planck Institute for Evolutionary 7 Anthropology, Deutscher Platz 6, 04103 Leipzig, Germany 8 9 Correspondence adress: [email protected] 0 10 Delimitation of Call Types of Red Crossbill (Loxia curvirostra) in the Western Palearctic 11 1 Abstract 12 13 Vocal signals are important in many animal species for communication, coordination, and 14 pair bonding and are especially well studied in birds. In the Red Crossbill (Loxia 15 curvirostra) calls are an important trait for mate choice. In this species, calls having the 16 same function (e.g., flight calls or excitement calls) are known to be clustered in distinct 17 groups, so called 'call types'. Each individual utters only calls of one call type. The driving 18 force for the differentiation of Red Crossbill call types in the Palearctic remains unknown, 19 as call types often overlap in space, and time and birds can be seen feeding on the same 20 seeds. In this study, we investigated calls of crossbills, recorded within seven years in the 21 Western Palearctic. We found at least 17 distinct call types of Red Crossbill and at least 22 two call types of Parrot Crossbill (Loxia pytyopsittacus). There were obvious differences 23 in call type delimitation between the northern and southern part of the study area. We 24 argue this is in conflict with the ecological differentiation hypothesis and propose that 25 there are other or further driving forces for this differentiation process. 26 27 2 Keywords 28 29 Red Crossbill; Loxia; curvirostra; pytyopsittacus; Population delimitation; Call types 30 1 31 3 Introduction 32 33 Vocal signals are used by many different animals for inter- and intraspecific 34 communication and serve many different purposes such as coordination within groups 35 (Lemures (Ramanankirahina et al., 2016)), group cohesion (Lemures (Braune, Schmidt, 36 & Zimmermann, 2005)), individual recognition (dolphins (Sayigh et al., 1998)), and pair 37 bonding (budgerigars (Hile, Plummer, & Striedter, 2000)). One of the most vocal groups 38 of animals are birds. Their vocalizations are grouped into songs and different kinds of 39 calls (Catchpole & Slater, 2008). Songs are usually complex vocal signals, mainly used 40 by males to establish a territory and to attract females. Calls are usually short and simple 41 structured vocal signals used in various contexts like threat, alarm or flight. The 42 distinction between song and calls is not always unambiguous, as many calls can be more 43 complex, and there seems to be a continuum between the two categories (Catchpole & 44 Slater, 2008). Most vocal signals are thought to be innate in birds, but some vocal signals 45 are known to be learned by members of three bird orders: hummingbirds (Trochilidae), 46 parrots (Psittaciformes) and songbirds (Passeriformes) (Catchpole & Slater, 2008). Birds 47 learn vocalizations during a sensitive period in the first few months of life (closed ended 48 learners, e.g., the White-crowned Sparrow (Zonotrichia leucophrys) (Marler, 1971)) or 49 throughout their lifetime (open ended learners, e.g., the Pine Siskin (Carduelis pinus) 50 (Mundinger, 1979)) with intermediate forms in between those two learning systems. 51 While song learning is well studied (Nottebohm, 1971), (Konishi, 1965), (Thorpe, 1954), 52 call learning has gained less attention because calls have been thought to be mostly innate 53 (Marler, 1963). However Mundinger (1979), compared the calls of social pairs of 54 different finch species and concluded that call learning is widespread within Carduelinae, 55 a subfamily of Finches (Fringillidae). 2 56 The Red Crossbill (Loxia curvirostra) is one of these call learning Carduelinae species 57 (Sewall, 2011). Crossbills give different calls in different situations and two calls are 58 particularly striking: excitement calls (ECs, sometimes also called ‘toops’ (Nethersole- 59 Thompson, 1975)) and flight calls (FCs). ECs are mostly used when a perched bird is 60 agitated. Flight calls are the most commonly uttered calls and are probably best 61 considered contact calls used both in flight and when perched. The Red Crossbill occurs 62 in coniferous forests (Picea spp., Pinus spp., Larix spp., Tsuga spp. and Pseudotsuga sp.) 63 in the northern Hemisphere (Clement & Christie, 2017). Their occurrence and movements 64 are closely linked to the availability of conifer seeds as their main food (Newton, 1972). 65 South of about 44°N, coniferous forests are dominated by different species of Pine (San- 66 Miguel-Ayanz et al., 2016). There, seed production of many Pine species is fairly even 67 and reliable year to year, such as in Mountain Pine (Pinus mugo uncinata) (Clouet, 2000), 68 Scot’s Pine (Pinus sylvestris) (Harper, 1977), Black Pine (P. nigra) (Tapias et al., 2011; 69 own evaluation of ICP Forests (www.icp-forests.net)) and Aleppo Pine (P. halepensis) 70 (Tapias et al., 2011; own evaluation of ICP Forests (www.icp-forests.net)). Thus, in this 71 region Red Crossbills live and feed in these trees all year round (Senar et al., 1993) being 72 resident or moving only short distances (Alonso et. al., 2016; SEO BirdLife, 2018). North 73 of 44°N the main food resource of Red Crossbills are seeds of spruce (Picea spp., 74 especially Norway Spruce (P. abies)) and larch (Larix spp.) (Alps (Glutz von Blotzheim 75 & Bauer, 2001), Germany (Thies, 1996), Great Britain (Summers & Broome, 2012), 76 Netherlands (Bijlsma, 1994)). In contrast to many pine species, the number of seeds in 77 spruce and larch varies considerably from year to year (Thies, 1996). Therefore, crossbills 78 in these coniferous forests are forced to leave areas during a cone crop failure and cover 79 long distances when searching for coning trees. They are thought to be nomadic (Perrins 80 & Cramp, 1998). 3 81 Many bird species, especially those learning calls, show geographical variation in their 82 vocalisations. Close relatives of crossbills with a linkage to a geographical area such as 83 Chaffinch (Fringilla coelebs) (Bergmann et al., 1988), Pine Grosbeak (Pinicola 84 enucleator) (Adkisson, 1981), and Bullfinch (Pyrrhula pyrrhula) (Constantin & The 85 Sound Approach, 2006) display such clear spatial variation in their calls. Other nomadic 86 species, like Siskins (Carduelis spinus and C. pinus), seem to show flock specific calls 87 which birds change when they join a new flock (Mundinger, 1979). Calls of Red 88 Crossbills vary too. Weber (1972) described two ‘call dialects’ of Red Crossbills in the 89 same area in Germany. In fact, FCs and ECs are clustered in distinct groups, the so called 90 ‘call types’, which are recognizable by their distinct spectrograms (Groth, 1993a). The 91 call type of an individual is therefore recognizable by its distinctive FCs as well as its 92 ECs (individuals sharing the same FC type usually also share the same EC type (Summers 93 et. al., 2002)). Individuals probably keep the same call type for their entire life, at least 94 for several years (Keenan & Benkman, 2008). In fact, a change in the call type of an 95 individual has been rarely described, and it is only documented for the FCs (Keenan & 96 Benkman, 2008). Calls are important cues for mate choice of Red Crossbill (Snowberg 97 & Benkman, 2007); hence, birds with different call types show more or less pronounced 98 genetic differences (Parchman et al., 2016). There are ten call types described in Red 99 Crossbill for North America (Groth (1993b), Benkman et al. (2009a) and Irwin (2010)), 100 and seven to nine call types in central and northern Europe (Clouet & Joachim (1996), 101 Robb (2000), Summers et. al. (2002), Edelaar et al. (2004), and Constantine & The Sound 102 Approach (2006)). However, these European studies are mostly restricted to single 103 countries, and a large-scale analysis of crossbill calls is lacking in the Western Palearctic. 104 Actually, within one forest, birds with different call types can often be found breeding 105 next to each other (Robb, 2000) and feeding in the same trees (Summers et al., 2010). 4 106 This differentiation without obvious spatial or ecological differences in the Western 107 Palearctic makes Red Crossbills a unique study species within birds, raising the question 108 about the drivers for the differentiation process. Edelaar et al. (2012) analysed crossbills 109 from Spain and found differences in morphology, genetics, and vocalizations between 110 crossbills foraging in different tree species and therefore argued, that differentiation of 111 crossbill calls is an effect of isolation by ecology. 112 In our study, we investigated call variation in crossbills throughout the Western 113 Palearctic. We clustered calls in groups, and tried to connect them with the different 114 species of Crossbills and the call types of Red Crossbill which had been published in 115 Europe before. If similar ecological preferences shape call types, we expect to find calls 116 to be clearly clustered in distinct call types all over the study area as result of a possible 117 speciation process. We expect only differences in call type delimitation between northern 118 (nomadic) and southern (resident) birds, if geographic origin promotes call type 119 differentiation due to the different movement patterns. 120 121 4 Material and Methods 122 4.1 Recordings and Classification 123 To analyse variation and number of call types of European Red Crossbill, we collected 124 8,216 recordings across the Western Palearctic (west of 60°E and north of 25°N) recorded 125 between June 2010 and May 2016. The recordings were made in 33 countries (Table S 126 1), and we acquired them from different sources (Table S 2).