Here we report significant differences between loud-call structures of P.thomasi, P.potenziani, P.comata and P.melalophos. Among the latter species a significant separation between the South Sumatran P.m.mitrata and the central Sumatran P.m.mitrata, as well as a further separation between P.m.melalophos from Bengkulu and the remaining P.m.melalophos populations, could be detected. The acoustic discrimination between Presbytis taxa was highly positively correlated with their genetic distance. In addition, we found substantial significant correlations between acoustic similarity and geographic distance and between genetic distance and geographic distance.
In our recent molecular genetic study  we suggested a paraphyly for P.m.mitrata, with the central Sumatran populations being closely related to P.m.melalophos and the South Sumatran populations forming a sister lineage to P.comata (Figure 5B). Our current findings on the acoustic structure of loud-calls strongly support these results.
P.m.mitrata was reported to inhabit the area southeast of the Batang Hari river, a large river in central Sumatra. In the west, this subspecies does not extend to the Bukit Barisan range, a mountain range on the western side of Sumatra , where P.m.melalophos occurs . Our samples of the central Sumatran P.m.mitrata (population 13) derived from the above described northernmost distribution range of this subspecies, south of the Batang Hari river. Although much paler, the morphological appearance resembles the reddish P.m.melalophos more than the grayish white southern Sumatran P.m.mitrata (Additional File 2). Whether there might be a transition zone between P.m.melalophos and P.m.mitrata demands further research. It is highly likely that P.m.melalophos gradually intergrades with P.m.mitrata, as may be the case between P.m.bicolor and P.m.melalophos . Our results, however, let us conclude that the central Sumatran P.m.mitrata population is the paler color variant of P.m.melalophos. Thus, the geographical distribution range of P.m.melalophos should be extended from the Bukit Barisan range eastwards towards Jambi. The southern Sumatran P.m.mitrata is genetically, morphologically and acoustically distinct from the remaining P.melalophos subspecies (see also Additional File 2). Therefore, if the Phylogenetic Species Concept [37, 38] is applied, P.m.mitrata would be elevated to a monotypic species P.mitrata Eschscholtz, 1821 .
Among P.m.melalophos we found the calls from Bengkulu (population 7) forming a distinct cluster. Unfortunately, genetic data from Bengkulu are lacking, but acoustically, the call types were more closely related to the Southern P.m.mitrata mainly due to the presence of inhalation elements. Historically different color morphs of P.m.melalophos were described, all of which are currently classified as synonyms of P.m.melalophos . These are a) the much less red and buffer variant from Bengkulu (Simia melalophos Raffles, 1821; syn. flavimanus Geoffroy, 1830), b) a foxy red northern form (Presbytis nobilis Gray, 1842) from Solok , c) a less reddish form from Padang (Semnopithecus ferruginneus Schlegel, 1876) and d) a golden buff variant (Semnopithecus sumatranus var. aurata Müller & Schlegel, 1841) from Gunung Talamau (ca. 150 km northwards from Padang) . The great diversity of color morphs in Presbytis, in particular in P.melalophos, has caused much debate over the past decades. Coloration might indicate relatedness, but can often be misleading, in particular, when no broad geographic sampling is available. Our data point out that the taxonomic ranking of some of these historically described taxa possibly should be reconsidered. However, the loud-calls from population 7 are only derived from two individuals and genetic data are missing. Therefore, further molecular genetic and bio-acoustic research based on a broader sampling is needed to draw final conclusions. Of great interest are the acoustic data of the Bornean taxa, in particular data of P.rubicunda. Based on molecular genetic results P.melalophos is even polyphyletic since P.rubicunda is nested within the P.m.sumatrana, P.m.bicolor, P.m.melalophos/central Sumatran P.m.mitrata clade . Previous studies already proposed a close affiliation of P.rubicunda and P.melalophos based on the red coat coloration  or in some aspects of behavior and vocalization . If species status of P.rubicunda is retained, species status of P.m.sumatrana, P.m.bicolor, P.m.melalophos will be consequently warranted, otherwise P.rubicunda has to be assigned as a subspecies of P.melalophos.
The correlation between acoustic structure and genetic differences was higher than the correlation between acoustic structure and geographic distance. This pattern can be explained by the following proposed Presbytis migration pattern, which is largely in agreement with Wilson and Wilson . The initial split in Presbytis occurred between P.thomasi and all other taxa, and P.thomasi colonized North Sumatra, which became isolated afterwards. The ancestor of the remaining taxa colonized first Borneo and later Sumatra. An early divergence of Bornean taxa is also supported by previous genetic studies [24, 32–34]. Of the ancestral Sumatran stock, one lineage invaded the Mentawai islands (P.potenziani), the other split into the proto-P.melalophos lineage and into the southern P.m.mitrata/P.comata lineage (Figure 5). Although calls from P.femoralis/P.siamensis (eastern Sumatra, Asian mainland) are not analysed in our study, previous publications show similarities in call structures of P.femoralis and P.thomasi [40, 41]. Our genetic study  shows that P.femoralis diverged relatively early from other lineages and, thus, the similar call structure of P.femoralis and P.thomasi might be a plesiomorphic feature. Up to this point the genetic, geographic and acoustic differences between populations increased. From this point onwards the geographic distances between populations decreased, because proto-P.melalophos subsequently transmuted into various present day subspecies, which were finally distributed across Sumatra as far as to the distribution range of P.thomasi in North Sumatra. Consequently, the geographic distance between P.thomasi and the remaining Sumatran populations decreased, while the genetic and the acoustic differences increased. Finally, the southern P.m.mitrata/P.comata lineage split into P.m.mitrata and P.comata that colonized Java. In this case we have a linear migration pattern and thus would expect a similar high correlation between acoustic structure, genetic and geographic distance, as it was currently shown in crested gibbons which are proposed to migrate in a linear fashion from North to South .
Surilis and gibbons are limited to rainforest habitats where the selection pressure forces an optimal adaptation of the structure of loud-calls for transmission over longer distances [5, 42]. Since the structure of loud-calls is inherited and call adaptation forces a similar structure, gene flow could achieve the major influence on the structural variation of calls . By combining the phylogenetic reconstruction of Meyer and colleagues  and the results of our study (Figures 2, 5A), we can observe a trend to simplification in call structure over time. However, it is difficult to explain why we found such a simplification in call structure. We cannot answer whether this is a general rule or whether this is a Presbytis-specific trait. Crested gibbons show an ambiguous result , where after a long period of syllable types with simple frequency modulation, a trend to a slightly more complex modulation appears. More acoustic comparisons with more species and at a higher taxonomic level are necessary to answer this question.
Interestingly, P.potenziani was regarded as most basal lineage  and due to similarities in call structure, the species was proposed as closely affiliated with P.thomasi . However, neither is the case, since P.potenziani derived much later . The specific call structure of P.potenziani is therefore either the result of an analogous evolution or a pleisiomorphic Presbytis feature. To clarify this issue further research is needed and particularly genetic and acoustic data on the Bornean and Malaysian taxa will help to better understand the evolution and phylogeography of the genus.
For instance, the call structure of P.rubicunda seems to be similar to P.melalophos calls  and, as discussed above, molecular genetic data also group P.rubicunda with P.melalophos . This close relationship can partly help to explain the interesting feature of general allopatry of respective Presbytis taxa in Sumatra, and sympatry in Borneo . P.rubicunda originated on Sumatra and subsequently invaded Borneo during the middle Pleistocene via a proposed connection between both islands . At this time Borneo was already colonized by the Bornean species P.chrysomelas, P.frontata and P.hosei. As a result of this second colonization, P.rubicunda is sympatric today with the three other species wherever their ranges overlap .