Evolutionary distinctness of Important Bird Areas ( IBAs ) of Sri Lanka : Do the species-rich wet zone forests safeguard Sri Lanka ’ s genetic heritage ?

Different components of biodiversity cannot be treated equally due to the limitations of knowledge and resources, and the heterogeneity in the distribution of biodiversity in the landscape. Therefore, biodiversity should be prioritized in effective sitebased conservation. Parameters related to ecology, sociology and economics are primarily used for this prioritization process. Genetic and evolutionary history is broadly ignored. Using Important Bird Areas (areas of high avian diversity or density, defined based on several key criteria of species richness and abandons) as a model, here we attempted to use several key evolutionary tools in biodiversity prioritization with the aim of testing their applicability in an island nation where evolutionary relationships of species are yet to be fully appreciated. We developed a DNA-based phylogenetic tree for the 342 total species of birds recorded in Sri Lanka’s 71 IBAs. We used DNA sequence archives at www.birdtree.org for closely-related crown nodes of Sri Lankan species and higher-order phylogenetic backbone for represented taxonomic orders to construct the phylogenetic tree. Using this tree, we calculated the Phylogenetic Diversity (PD), Evolutionary Distinctness (ED) and the extinction risk of deep lineages (EDGE scores) for each IBA. Batrachostomus moniliger (Ceylon Frogmouth) is the most ED species while Otus thilohoffmanni (Serendib Scopes-owl) is the most EDGE species for Sri Lanka. Sri Lanka’s premier dry zone National Parks, Yala, Bundala and Udawalawe are the top ranked IBAs in PD, ED and EDGE scores. Except Sinharaja MAB Reserve and Knuckles, all other wet zone IBAs, which are characterized by their high endemicity and biodiversity were not ranked among the top ten IBAs for above evolutionary parameters. Mixing of both continental and insular lineages in the dry zone due to its proximity to mainland, the presence of wetlands that support migratory lineages and the movement of wet zone taxa along the riverine forests towards the dry zone could have increased the PD and ED scores in dry zone IBAs. However, wet zone IBAs are better at keeping the avian genetic composition that is unique to Sri Lanka by holding most of the avian endemics. For islands like Sri Lanka, global indices do not reflect the local evolutionary and conservation status. Effective site-based conservation programs that are aimed at preventing local extinctions therefore, must recognize the heterogeneity of biodiversity across the landscape. Our results show that the evolutionary parameters such as PD, ED and EDGE score are effective tools to refine the conventional indices used in habitat prioritization for Sri Lanka.


INTRODUCTION
As a result of excessive consumption of natural resources and the climate change, world is experiencing a sharp drop in biodiversity and an unprecedented rate of extinction of both fauna and flora (Vitousek et al., 1997).Conservation of species is therefore paramount in curtailing biodiversity decay.With limited amount of knowledge and resources available, effective conservation requires prioritization of biological entities such as the species and ecosystems.Commonly used ecological parameters such as species richness and number of endemic species do not consider the evolutionary history of the species.Not all species are ecologically equally important, yet they are important in other aspects such as form, function, evolutionary uniqueness and aesthetic appeal, where an extinction of such species causes a considerable loss of biodiversity because biodiversity is not only species richness but also the history of life as a whole (Jetz et al., 2014).
The evolutionary history of a clade (a distinct group of organisms) can be traced back using a phylogenetic tree (Pagel, 1999), in which the tips and nodes of the tree stand for descendant taxa and their common ancestors respectively (Vellend et al., 2011).Nee and May (1997) suggested that the more distal nodes (i.e.species) a branch carries, the lower the amount of evolutionary history captured by each of its representative nodes in that particular part of the tree of life (Figure 1).

Several phylogenetic parameters have been introduced
to measure how much evolutionary history is captured by a species or a geographic area such as: Phylogenetic Diversity (PD), Evolutionary Distinctness (ED), and Evolutionary Distinct and Globally Endangered Score (EDGE Score) (Faith, 1992a;Isaac et al., 2007;Vellend et al., 2007;Jetz et al., 2014).PD is the minimum sum of branch length of a particular set of taxa in the phylogenetic tree (Faith, 1992a).The branch length between two given nodes (i.e.taxa) is the estimate of time at which two nodes started to evolve independently of each other (Vellend et al., 2011).ED is a measure of how much a particular species is isolated in a phylogenetic tree.The higher the ED of a given species, the lower the number of closely related species it has in the phylogenetic tree (Jetz et al., 2014).The EDGE score (Isaac et al., 2007) is an index derived from combining ED scores with the IUCN red list categories (Butchart et al., 2004).EDGE scores can Ceylon Journal of Science 46 (Special Issue) 2017: 89-99 be directly used to set conservation priorities (Isaac et al., 2007).
Mean Pairwise Distance (MPD) is another phylogenetic measurement that is used to determine whether the species found in a given area are more closely related than predicted by their form and function.MPD is measured as a summation of the evolutionary relatedness of all the pairs of species found in that area (Webb et al., 2002).
Effective conservation could be achieved through sitebased conservation measures (Jetz et al., 2014).Since the global distribution and abundance of taxa is not uniform (Brusatte et al. 2015, Jetz et al. 2014), similar to what has been recognized in IUCN red listing criteria (National Red List, 2012), the conservation prioritization should recognize the heterogeneity of evolutionary and ecological criteria across the landscape.Therefore, global PD, ED or EDGE values do not fit to assess the level of accumulation of lineages and risk of local extinctions of globally widespread species in the regional or national level (e.g. in an island like Sri Lanka).There are numerous criteria available for site selection.For example biodiversity indices such as Simpson Index (Simpson, 1949), legislative descriptions such as National Parks and sites marked for species based conservation such as 'Important Bird Areas' (IBAs) can be used to compare sites.
IBAs are sites that have international significance on the conservation of birds and were introduced by a global consortium of bird-related organizations called BirdLife International (BirdLife International, 2016).IBAs have both regional and global significance; they have been selected based on scientific and objective standards associated with globally threatened species, restrictedrange species, biome-restricted species and large bird aggregations (BirdLife International, 2016).There are 71 IBAs in Sri Lanka and all of them are located within the protected area network of Sri Lanka (Field ornithology group of Sri Lanka, 2003).
Birds have evolved within the lineage led to dinosaurs over a period of ~100 million years (Brusatte et al., 2015).About the past 50MYA they underwent an explosive radiation which led to the present diversity of ~10,000 species (Jarvis et al., 2014, Jetz et al., 2012).Does each of these extant species capture the same evolutionary history in the bird tree?Do all IBAs carry similar evolutionary history of the birds that they harbor?Which IBAs have higher evolutionary significance or carry a higher number of unique or endangered lineages, and hence require greater conservation attention?Using IBAs as a model, here we attempted to use several key evolutionary tools such as PD, ED and EDGE, to rank IBAs in the aim of testing their applicability as a tool for conservation for Sri Lanka.We hypothesized that the species-rich wet zone IBAs have greater PD, ED and EDGE scores compared to their less celebrated dry zone counterparts.We use the help of lowcost global data archives and statistical freeware to perform our bioinformatics analysis.

METHODS
A total species list, which contains all recorded species across 71 IBAs was obtained from the IBA database of the Field Ornithology Group of Sri Lanka.Vagrant species and species unlikely to utilize the focal habitat (e.g.seabirds in montane IBAs) were removed from the dataset.The final list of 342 species was fed into the option 'Phylogeny subsets' of 'www.birdtree.org'and a set of 5000 phylogenetic trees was generated.Hackett et al. (2008) described the deep order-and family-level affinities of birds.We used the option 'Hackett All Species: a set of 10,000 trees with 9,993 OTUs each' in the www.birdtree.org site, which uses the database of species of Hackett et al. (2008) as the backbone.To generate the grand tree for all 342 species recorded, the program followed four main steps.For the construction of the tree it combined relaxed molecular clock trees of well supported clades with a backbone which contained representatives from each clade (Jetz et al., 2012).As the first step, each species was assigned to one of the 158 clades which arise from the 158  (Nee & May, 1997).
tips of the backbone trees.Then relaxed-clock trees were constructed for each clade.The species that do not have genetic information were plugged in to their relevant clades using the information obtained by combining the relaxed clock trees and available taxonomic information (Jetz et al., 2012).The final tree was constructed by merging trees from either of the previous two steps with time calibrated relaxed molecular clock backbone trees (Jetz et al., 2012;BirdLife International, 2016).
A consensus tree (Figure 2) was generated from the file of 5,000 phylogenetic trees generated by www.birdtree.org using 'TreeAnnotator v1.8.2' of the software BEAST (BEAST v1.8.2).R platform (R Development Core Team) was used to measure the Evolutionary Distinctiveness (ED) and EDGE scores for each species.Using the R codes E1 (see below), the ED of all 342 species was calculated.
Originality<-evol.distinct(tree,type="fair.proportion" (E1) The ED of IBAs was calculated using the equation E2, ED of an IBA= ∑ ED of each species in the IBA (E2) EDGE score was calculated for each species using E3 (Isaac et al., 2007)
Then the EDGE scores for each of the IBAs were calculated separately by summing the EDGE scores of all the bird species in the IBA (E4).EDGE score of an IBA= ∑ EDGE of all species in the IBA (E4) We ranked all 342 species based on their ED and EDGE values.The global ED and EDGE scores of birds were taken from Jetz et al. (2014).The MPD could not be calculated due to several technical limitations.
Species Richness (SR) and abundance are two of the most widely used ecological parameters in the measurement of biodiversity (Gugerli et al., 2008).Number of endemics in a given area is another parameter that can be used to Dilini K. Abeyarama and Sampath S. Seneviratne measure biodiversity.SR of each IBA was obtained taking the total number of species in the total area of each IBA.Number of endemic species (EM) of an IBA was obtained by counting the total number of endemic species of birds found in each IBA.
When considering both the resident and migratory species, the below 10 IBAs topped the list of IBAs with highest PD; Yala National Park, Bundala NP, Udawalawe NP, Kalametiya Sanctuary, Wasgomuwa NP, Sigiriya Sanctuary, Wilpattu NP, Gal Oya NP, Minneriya-Girithale NP and Maduru Oya NP (Figure 3).When only the resident species were included for the analysis, the below IBAs topped the list of IBAs with the highest PD; Yala NP, Udawalawe NP, Wasgomuwa NP, Bundala NP, Gal Oya NP, Sigiriya Sanctuary, Wilpattu NP, Maduru Oya NP, Minneriya-Girithale NP and Sinharaja MAB Reserve.Therefore, Sinharaja MAB Reserve is the only wet zone IBA in the list of top IBAs with highest PD (Table 03 and  Table 04).
In the analyses with migratory species and without migratory species, a higher percentage of IBAs that ranked top 10 under four parameters SR, PD, ED and EDGE score are forest IBAs located in the dry zone (Figure 3).However, when considering the number of endemics per IBA, the top 10 IBAs are all forest IBAs in the wet zone (Table 05-07).

DISCUSSION
Species in the older and species-poor lineages tend to carry a greater amount of phylogenetic history than those in speciose, recently branched out lineages (Isaac et al., 2007).Using several phylogenetic parameters such as Phylogenetic Diversity (PD), Evolutionary Distinctness (ED) and Evolutionary Distinct Endangered Lineage (EDGE) scores, we measured how much evolutionary history is captured by Sri Lanka's IBAs.We tested whether the avian evolutionary history of the island is confined to a particular climatic zone.When we ranked IBAs based on PD, ED and EDGE scores the majority of the top 10 ranked IBAs are located in the dry zone (Fig. 03).Although wet zone IBAs are poor at representing the evolutionary history (PD and ED) of birds of the island, their high endemicity represents the genetic heritage unique to the island.
The PD of an IBA increases with the increment of the number of species in that IBA with some distinguished omissions.For example, the SR of two IBAs Anuradhapura Sanctuary and Sinharaja MABR are 172 and 182 where the PD of them are 5492.97and 5241.96respectively.The reason for this is that not all the species contribute equally for PD (Brusatte et al., 2015).The species that have evolved long ago have greater contribution to PD while the recently diverged species have lesser contribution (Hackett et al., 2008; Table 01).The species that have low ED scores are not isolated in the avian phylogenetic tree and they have many closely related species.However the birds that have very high ED scores such as B. moniliger (Ceylon Frogmouth) are isolated groups in the phylogeny and they do not have closely related forms near them in Sri Lanka.The species such as S. bengalensis (Lesser-Crested Tern) and S. bergii (Great-Crested Tern) that have similar ED scores are sister to each other.Closely related (nonsister) forms could also have approximately similar ED scores (Table 01).
The cumulative ED score of an IBA represents how much evolutionary history is captured by that particular area (Jetz et al., 2014).The higher the species richness, the higher will be the ED captured by that area.When that IBA is home for more evolutionarily isolated species (species with high ED) the ED value of the IBA could increase more than the average.For an example, the two IBAs Newgalkanda and Padaviya Tank harbor 66 species in each, but the ED are very different between the two IBAs (1464.891and 1739.62 respectively).Padaviya Tank IBA must be carrying more evolutionary isolated species than that by the Newgalkanda IBA.
Yala NP is rank 1 st in SR, PD, ED and EDGE score in both analyses done with and without migratory species (Table 05-07).The reason could be its very high SR.Species richness tends to push PD and adds a bias.Using MPD one could reduce this bias.However due to technical difficulties, we could not correct this bias.When only resident species were included in the analysis, the order of IBAs with regards to PD, changes.In our study at Bundala NP, which ranked 2 nd when migratory species were included, ranked 4 th when they were removed.Being a wetland IBA and a key overwintering location for migratory species, the reason for this shift is that Bundala NP gets several extra lineages as migratory species, which elevates its PD.
The local and global ED ranking are very different, for example, P. brachyura (Indian Pitta) which ranks 3 rd in the local rankings comes 10 th if we use global rankings.But P. haliaetus (Osprey) which ranks 8 th under local rankings will come 1 st with the global rankings.The reason for this is that in the local context Indian pitta represents a monotypic genus and therefore its ED is high, but when all bird species of the world are considered the pittas represent a polytypic genus.
Sri Lanka is a distinctly identifiable geological entity (a continental island) with a unique geological history.As a result, it harbors a unique faunal and floral assemblage (e.g.Ripley and Beehler 1990, Bossuyt et al 2004, Wickramasinghe et al 2017).Even though there are ~10,000 species of birds in the world, the island of Sri Lanka only carries a 1/20 th of the global diversity that is ~ 500 species (Rasmussen and Anderton, 2005).To evaluate the evolutionarily significance of Sri Lanka's avifauna, one should only consider the 500 or so species found in Sri Lanka.To simplify the concept, let us breakdown this approach into few simple steps: (a.)Let us go to Sri Lanka in time T 0 where no birds found.(b.)In time T 1 one species  With that the ED of the SP1 drops by half.(d.)With the colonization of the SP3 in T 3 the PD improves further and ED of each of the three species reduced.Regarding the phylogenetic position of SP3 the reduction of ED will vary in the three species (Isaac et al., 2007;Vellend et al., 2007).(e.)After T n (say today) there are 500 species of birds found in Sri Lanka.Those birds represent the total phylogenetic diversity of birds in the island and each one of the 500 species will have its own ED for the area -that is the fraction of evolutionary information that the species is represented.Bird lineages that are represented by few species will have high ED, while the members of the more speciose lineages will have low ED for the area.Note that still there are nearly 10,000 species of birds in the rest of the world.Since they are not present in Sri Lanka, they do not contribute to either of the tested parameters.
No matter how speciose a lineage is in the global arena, the species' evolutionary value depends on the number of members represented by that lineage in the focal geographic area.Therefore, the evolutionary weightage of species varies from region to region.Consequently, to assess the evolutionary distinctness of Sri Lankan birds found in IBAs one should not consider species that are not found in the island or away from IBAs.We want to see the phylogenetic information that each species brings in to these IBAs.This approach is similar to the IUCN Red list approach.The red list has two independent versions: the Global Red List and the National Red List (National Red List 2012).
For the national list one should only use the species found in the focal area (i.e.Sri Lanka).A globally least concern (LC) species can be critically endangered (CR) in the national list.Similarly, globally endangered species (EN) could be LC in the national list.The national status is evaluated independent of the global status (National Red List 2012).
We believe that for effective site-based conservation and management, one should honour the heterogeneity of these measures across the landscape.In order to prevent local extinctions of globally widespread species, especially in small islands, conservation efforts should focused on the regional and local diversity irrelevant to the global figures.
The EDGE score of a species depends on both the ED and the IUCN red list status (Isaac et al., 2007).For a species to have a high EDGE score, that species should be an evolutionary isolated species, a globally threatened species or both.O. thilohoffmanni (Serendib Scops-owl) has the highest ED for Sri Lanka, it has a relatively low ED score.But it is an endangered species.The EDGE score for B. moniliger, which has the highest ED value for the country, has a low EDGE score due to its better IUCN red list status (Table 02).None of the top 10 ranked species under ED is endemic to Sri Lanka; however, the majority of the top 10 EDGE species are endemic (Tables 01-02).Most of our endemic forms are diverged recently as insular populations (Rasmussen and Anderton, 2005;Ripley and Beehler, 1990) hence low ED, and that most of them have poor red list status (National Red List 2012) hence the higher EDGE score.
IBAs in Sri Lanka are distributed in three main climatic zones as wet zone, dry zone and arid zone (Fig. 03).Top IBAs with highest SR are located in dry zone.Just two top-ranked IBA are found in the wet zone (Table 05-06).None of the IBAs out of the 10 IBAs with highest PD are located in the wet zone either (Table 05-06).Top 10 IBAs under ED and EDGE scores are also dominated by dry zone IBAs (Fig. 03).Only Sinharaja MABR and Knuckles CR represented the wet zone under these parameters.However, when the number of endemics were considered, the opposite is true (Table 05-06).In general, the sizes of the IBAs in dry zone are bigger than that of the IBAs in wet zone (Fig. 03: unpublished data -FOGSL IBA database).Because of that higher carrying capacity of dry zone IBAs, they carry slightly higher number of species (SR).Furthermore, most of the IBAs in the coast (Figure 3) receive migratory waterbirds, which increase the SR of these IBAs as well.
Most of these lineage-rich dry zone IBAs are fed by rivers that are originated from the wet zone (Figure 3).Allowing the mixing of wet zone (hydrophilic) lineages with the dry zone (hydrophobic) lineages, which makes the riverine forests of the dry zone harbor an assortment of both wet and dry zone taxa.The high PD, ED and SR observed in these IBAs could be a result of this lineage-rich riverine forests.Although wet zone IBAs of Sri Lanka are not best at harboring the evolutionary history of birds, they are the bests in up keeping the avian endemics and EDGE species of the country.

Figure 1 :
Figure1: A schematic phylogenetic tree illustrating the unequal representation of evolutionary history captured by each of the nine distal nodes (A to A'') after(Nee & May, 1997).

Figure 2 :
Figure 2: Phylogenetic tree of total 342 bird species reported in IBAs of Sri Lanka.

Figure 3 :
Figure 3: IBAs of Sri Lanka.The top five most evolutionarily diverse (PD, ED and EDGE) IBAs are highlighted in red color.Main climatic zones and main river systems of the island are also indicated in the map.

Table 01 :
Species that has the highest evolutionary distinctness (ED) in Sri Lanka

Table 02 :
Species that has the highest EDGE score in Sri Lanka

Table 03 :
Top 10 IBAs with highest phylogenetic diversity (PD).Both resident and migratory species of birds were included for this analysis.

Table 04 :
Top 10 IBAs with highest phylogenetic diversity (PD).Only resident species of birds were included for this analysis.

Table 05 :
IBAs with highest scores in SR, EM, PD, ED and EDGE scores with respect to the climatic zone.Both resident and migratory species are included in the analysis.Colour key: Tangerine-arid zone, coral-dry zone, dark green-wet zone hill country, and green-wet zone low country.

Table 06 :
IBAs with highest scores in SR, PD, ED of IBA and EDGE scores with respect to climatic zones.Only the resident species are included in the analysis.Colour key: Tangerine-arid zone, coraldry zone, dark green-wet zone hill country and green-wet zone mid & low country.

Table 07 :
IBAs with highest scores in SR, EM, PD, ED and EDGE scores with respect to the habitat type (forest or wetland).Both resident and migratory species are included in the analysis.Colour key: Green-forest IBAs, blue-wetland IBAs and cyan-forest and wetland IBAs.SP1) had colonized in Sri Lanka.Therefore, now the evolutionary history (PD) of birds in this island is represented by SP1.There are nearly 10,000 species of birds found in the rest of the world, however none represented in the focal area, therefore the SP1 solely represents the bird lineage in Sri Lanka at T 1 .As a result ED of SP1 should be very high.(c.)In T 2 another species of bird (SP2) got colonized in the island.Now the total PD is represented by SP1 and the extra phylogenetic info brought in by SP2.