Molecular identification of Haemosporidia in avian endemics of Gorgona Island within a context for the eastern tropical Pacific region

Journal Pre-proof Molecular identification of Haemosporidia in avian endemics of Gorgona Island within a context for the eastern tropical Pacific region

Raul Sedano-Cruz, Andres Castillo, Diana Lorena Gil-Vargas PII:

S1567-1348(19)30349-1

DOI:

https://doi.org/10.1016/j.meegid.2019.104123

Reference:

MEEGID 104123

To appear in:

Infection, Genetics and Evolution

Received date:

14 August 2019

Revised date:

31 October 2019

Accepted date:

16 November 2019

Please cite this article as: R. Sedano-Cruz, A. Castillo and D.L. Gil-Vargas, Molecular identification of Haemosporidia in avian endemics of Gorgona Island within a context for the eastern tropical Pacific region, Infection, Genetics and Evolution(2018), https://doi.org/10.1016/j.meegid.2019.104123

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© 2018 Published by Elsevier.

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Molecular identification of Haemosporidia in avian endemics of Gorgona Island within a context for the Eastern Tropical Pacific region

Raul Sedano-Cruz1,* , Andres Castillo1 , Diana Lorena Gil-Vargas 1 Universidad del Valle 1Grupo de Ecología Animal, Department of Biology, Universidad del Valle, Calle 13 No 100-00, Edif. E20, Office 3120, Cali, Colombia.

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3 [email protected]

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4 [email protected]

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Running Head: Avian Haemoproteus on Gorgona Island

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*Corresponding author: Universidad del Valle, Calle 13 No 100-00, Edif. E20, Office 3120, Cali, Colombia. Tel.: (+57 2) 321 21 00, Ext. 3241.

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E-mail address: 2 [email protected] (R. Sedano-Cruz)

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ABSTRACT Island bird populations and their obligate blood parasites are of interest for understanding the accumulation of biodiversity and the evolutionary relationship with their mainland congeners. We examined avian Haemosporidia cytochrome b gene among terrestrial birds on Gorgona Island National Park, Colombia. Three Haemoproteus haplotype groups found on Gorgona Island have a higher genetic similarity to Haemoproteus found in the eastern tropical Pacific than

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those documented in Africa, Asia, Europe and Oceania. Two of the haplotype groups on the

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island are generalists in terms of infecting multiple hosts and their wide geographical distribution

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within the eastern tropical Pacific region, a third Haemoproteus haplogroup appears endemic to

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Gorgona Island. The overall prevalence of haemosporidian parasites is 57,9% for birds on

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Gorgona island, which is higher than local reports of prevalence documented in other archipelagos or the mainland. The island population of Cyanerpes cyaneus gigas seems to be the

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most susceptible to Haemoproteus infection when compared to Thamnophilus atrinucha gorgonae and Coereba flaveola gorgonae. Our findings support a ubiquitous pattern of

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endemism among organisms including avian haemosporidian parasites on Gorgona Island and

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also highlight the potential exposure of island bird populations to avian malaria.

Keywords: Haemosporidia, Island bird populations, parasite-host, Gorgona Island, malaria.

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1. Introduction Avian haemosporidians are worldwide obligate blood parasites, these protozoa belong to the genera Leucozytozoon, Plasmodium and Haemoproteus (Levine, 2018). Avian malaria has been shown associated with the demise of island bird populations, as in New Zealand (Alley et al., 2008; Niebuhr et al., 2016) and Hawaii (García-Longoria et al., 2015). In the Galapagos Islands

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reservoir species of avian haemasporidians have been documented (Levin et al., 2013, 2009), but

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a negative effect on bird species has not been clearly established (García-Longoria et al., 2015). In contrast, in the islands of the Caribbean basin the presence of avian malarial parasites further

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indicates that resident birds seem somewhat more susceptible to malaria than the introduced bird

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species in the islands of the Antilles (Atkinson et al., 2000; García-Longoria et al., 2015). This

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further suggests that some avian haemosporidians may constitute a threat to insular populations (Valkiūnas, 2005). Therefore, monitoring haemosporidians in island bird populations is

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indispensable to determine conservation actions.

In islands, avian parasites are constrained to a more limited array of hosts since islands only harbour a reduced segment of the mainland’s terrestrial biodiversity. Since the diversity of parasites in a habitat tends to increase with an increased diversity of hosts (Gil-Vargas and Sedano-Cruz, 2019; Kamiya et al., 2014), one would expect that the more limited host opportunities in islands may result in a community of generalist parasites (Fallon et al., 2003). Therefore, the understanding of the distribution of parasites in islands can be of value to estimate the amplitude of host-parasite interactions (Apanius et al., 2000).

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Although the genera Plasmodium (Beadell et al., 2006; Ewen et al., 2012; Tompkins and Gleeson, 2006) and Haemoproteus (Apanius et al., 2000; Fallon et al., 2005) tend to predominate in islands. Their pattern of colonization is far from well understood. Nevertheless, it has been documented that Haemoproteus is less host-specialized than Plasmodium in South America

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(Clark et al., 2014). It stands to reason that the introduction of Plasmodium or Haemoproteus to

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an island may be mediated by the availability of vectors, which diversity in turns, tends to be lower in islands as well. Thus, studying the spatial distribution of avian haemosporidians in

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islands is also important to understand the nuances of parasite-host relationships over large

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geographic breadth.

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Gorgona Island is located on the continental shelf of the Colombian Pacific coast. It is separated

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by 35 km of a maritime gap, and it is characterized by climatic conditions somewhat different from nearby areas on mainland (Blanco, 2009). Gorgona island is a National Park since 1986 and

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constitutes an important hotspot of biodiversity of the eastern tropical Pacific (ETP). This island

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houses several endemics including plants, invertebrates, reptiles, and mammals (Giraldo et al., 2014). The bird species on the island can be roughly grouped according to their ecological attributes in residents, migrants, marine and shorebirds to focus conservation strategies. But only 14 resident avian species have been reported (Giraldo and Garcés, 2012), from which three are poor flight island endemic populations (Thayer and Bangs, 1905). The understanding of how avian malaria circulates among bird fauna on this island remains to be examined yet.

Here we are documenting the prevalence and the variation of the avian Haemosporidia cyt b gene in birds of Gorgona island. The molecular detection of parasites has become an efficient 4

Journal Pre-proof method to determine the number of infected individuals and the heterogeneity in the prevalence of avian haemosporidians suggests variations of host-parasite associations among island populations (Apanius et al., 2000). Therefore, knowing the existence of differential prevalence of avian haemosporidians between islands and the mainland may have significant evolutionary implications for parasites and hosts (Beadell et al., 2007; Hart, 1990).

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Our first objective was to establish if prevalence is uniform among endemic bird populations to

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the island. Thus, we compared sequence data of Haemosporidia among avian populations of the island. Using a phylogenetic context, we examine the geographic distribution of parasites and

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hosts by using accessions from the GenBank (Benson et al., 2011) and MalAvi (Bensch et al.,

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2009). In addition, we compare the genetic structure between avian malaria found on Gorgona

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Island among several regions of the world. Finally, considering that Gorgona Island exhibits high levels of endemism of flora and fauna (Giraldo and Garcés, 2012; Yockteng and Cavelier, 1998),

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we hypothesized the possibility of exclusive avian Haemosporidia to the island.

2. Materials and methods 2.1.

Study Area

Gorgona Island (2.9683° N, 78.1844° W) has a well-documented history of native americans, pirates and conquerors that used the island as a stopover. This little island of 26 Km2 also has a history on extensive habitat transformation due to livestock, overpopulation and the introduction of mesopredators over a complex phenomenon somewhat traceable since the XVI century (Giraldo et al., 2014). The weather is humid and the temperature stable averaging 26 ºC throughout the year (Blanco, 2009; Rangel-Ch., 1995). The annual rain (6.661 mm) doubles that 5

Journal Pre-proof of nearby locality 30 km away from the mainland and such an annual rain pattern is unimodal with positive net availability of water even in March, the driest month of the year on Gorgona Island (Blanco, 2009). The terrestrial habitat is under restoration since the island became a National Park in 1986.

2.2.

Sample collection, PCR Products, and DNA Sequencing

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Samples of bird species were collected between November 2016 and May 2018 in the Gorgona

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Island. We mist-netted birds and collected a ca.10 µL blood-drop from capillary brachial vein

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puncture and was preserved it in a lysis buffer (Longmire et al., 1992). The polymerase chain

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reaction has been instrumental to the rapid production of amplicons and sequencing of the mitochondrial cytochrome oxidase b gene (cyt b) of avian Haemosporidian parasites (Bensch et

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al., 2000; Hellgren et al., 2004; Lotta et al., 2019). The quality of the extracted DNA using the

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Thermo-Scientific® Kit was examined by PCR of host NADH dehydrogenase 2 mitochondrial DNA (mtDNA), following a previously described procedure (Sedano and Burns, 2010).

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Infections of malaria parasites were identified by PCR using 470 base pairs of the cyt b gene of

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avian Haemosporidia, including a negative control routinely. We used primers that have shown sensitivity to Leucozytozoon, Plasmodium and Haemoproteus (Harrigan et al., 2014), the nested PCR conditions were previously reported (Waldenström et al., 2004), with consistent efficiency (Chasar et al., 2009). The PCR products were visually confirmed in a 2% agarose gel using Safe DNA Dye (HydragreenT M), and such products were sequenced using BigDye (Applied BiosystemsT M). The DNA electropherograms were visually edited using Sequencher 4.1 (Gene Codes Corp.). This study was approved by the Universidad del Valle (Univalle) research ethics committee.

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2.3.

Prevalence and Haplogroups Identification

The total prevalence of Haemosporidia for Gorgona Island was calculated as the number of infected individuals divided by the total of the individuals sampled. Specific prevalence was calculated for those populations with five or more individuals sampled. The haplogroups of cyt b sequences from Gorgona Island were identified through clustering among 1686 sequences

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obtained from GenBank (Benson et al., 2011) and MalAvi (Bensch et al., 2009), using

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USEARCH v8.1 (Edgar, 2010) as implemented by Gil-Vargas and Sedano-Cruz (2019) through

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a centroid sequence, by a cut-off level of 99.3% of sequence identity to define a haplotype group.

Host Amplitude by Haemosporidian Haplogroups

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We examined the amplitude of hosts infected by Haemosporidian parasites found on the

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Gorgona Island and mainland. The amplitude of infected hosts is shown as the average number of avian species per genus; therefore, we calculated the avian species-to-genus ratio (S/G) by

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each haemosporidian haplotype group found on the island. The purpose of this calculation is to

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describe host S/G ratio beyond Gorgona Island for a given parasite haplogroup found on the island. The host S/G ratio higher than one shows that the number of known infected hosts species is higher in number of infected host genera by a given parasite haplogroup in the sample. In order to implement this calculation, we obtained host taxonomic data per parasite haplogroup from the additional file 3 from the data set of Gil-Vargas and Sedano-Cruz (2019).

2.5.

Analysis of Molecular Variance (AMOVA)

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Journal Pre-proof We calculated the number of variable sites of cyt b gene, heterozygosity, nucleotide diversity and Tajima´s D (Tajima, 1989). An AMOVA was implemented using ARLEQUIN 3.5 (Excoffier and Lischer, 2010) to examine the spatial genetic variation of avian Haemosporidia between that found in the Gorgona Island and several geographic regions as Europe, Asia, Africa, Oceania and including the ETP (see Appendix, Table B1). But within the ETP region, we examined the Haemosporidia spatial genetic structure using a model of isolation-by-distance (IBD) (Bohonak,

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2002), by implementing a Mantel test using a GTR substitution model (Tavaré, 1986) on genetic

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distances obtained with MEGA (Tamura et al., 2013). We further conducted a second AMOVA

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among groupings of hosts (resident, migratory, marine and shorebirds species). Although these

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groupings are not part of a natural categorization, they are important to the comparison of

Phylogenetics

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2.6.

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general ecological attributes of avian species.

We conducted a phylogenetic inference for haplotype groups found in the ETP using a

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GTR+I+G substitution model in BEAST v1.8.3 (Drummond et al., 2012), using the CIPRES 3

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platform (Miller et al., 2010) and a strict clock of 1.2% substitutions per million years (Ricklefs and Outlaw, 2010). The consensus tree was obtained using PAUP4 (Swofford, 2002), with a burn-in of ca. 25% of the trees output. Furthermore, the genealogy was visualized in ITOL using a node support ≥0.95 posterior probability (Letunic and Bork, 2007).

3. Results 3.1.

Parasite Prevalence

A total of 57 individuals of eight species were examined for the cyt b gene of avian

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Journal Pre-proof Haemosporidia and 33 individuals (57.9%) were identified as PCR positives (see Table 1 and Appendix, Table B2) and the average prevalence by bird species was 42.9%. This sample size for the small island area of 26 km2 is comparable with sampling sizes at the local scale of studies on haemosporidian in the Neotropics (Cadena Ortiz, 2015; Gonzalez-Quevedo et al., 2016; González et al., 2015; Harrigan et al., 2014; Jones et al., 2013). We obtained 21 partial sequences of Haemoproteus (MF990712-MF990716, MF990718-MF990729, MF990731-MF990734)

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further associated to three out of 78 Haemoproteus haplogroups found in the ETP (Fig. 1). The

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haplogroups found on Gorgona Island show lower diversity and the population is not yet in

3.2.

Haplotype groups distribution

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equilibrium according to the Tajima´s D test (Table 2).

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The H4, H39 , and H502 are Haemoproteus haplotype groups found on Gorgona Island. The latter is

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found exclusively on the island infecting Sporophila luctuosa and it is rare anywhere across the ETP (see Fig. 2). In contrast, H39 is the most common, infecting the majority of species in our

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sample (5/8), and 20 GenBank sequences are also represented within this haplotype group (see

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Appendix, Table B3). Both H39 and H502 share a most recent common ancestor in a clade of haplogroups infecting Passeriformes in Colombia (see Fig. 2 and Appendix, Table B3). The H4 Haemoproteus haplogroup infects exclusively C. flaveola in the Gorgona Island and it belongs to a clade with 19 sequences found in Colombia, Antilles, French Guiana, Brazil, Uruguay and the United States (see Appendix, Fig. 2 and Table B3). The divergence times estimate suggests that the aforementioned clades began to diversify in their haplotype groups around 0.53 and 0.21 million years before present, respectively (see Fig. 2).

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Journal Pre-proof 3.3.

Host Amplitude by Haemosporidian Haplogroups

The amplitude of infected hosts is higher for those parasites with a larger geographic breadth (see Appendix, Table B4). It is not surprising that H4 haplotype group found on Gorgona Island with the larger geographic amplitude also has the larger host S/G ratio (1.23). In contrast, H 39 and H502 haplotypes groups with a restricted distribution in Colombia have a low host S/G ratio (1.0). However, if we examine the host S/G ratio only for Gorgona Island, we found that in our

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sample the ratio is numerically equivalent for the three Haemoproteus haplotypes groups

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(S/G=1.0). This further suggests that given our sample at least H4 haplogroup is somewhat

Spatial Genetic Variation

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3.4.

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amplitude of hosts outside the Gorgona Island.

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constraint to a reduced breadth of hosts in the island as compare to what is its expected

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The comparison among regions worldwide suggest that Gorgona Island haplogroups are more differentiated from Haemoproteus genetic variation from Oceania (Φst =0.61153, P = 0.00001),

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Africa (Φst =0.57443, P = 0.00001), Asia (Φst =0.56705, P = 0.00001), Europe (Φst =0.52534, P (1, 234) =

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= 0.00001) and the ETP (Φst = 0.23663, P = 0.00001) (see Table 2, AMOVA Φst

0.25534, Exact Test P = 0.00001). This implies that Haemoproteus haplogroups found in bird species of Gorgona Island have a rather restricted circulation to the Americas.

Within the ETP, we found a differential distribution of avian Haemoproteus genetic variation between Gorgona Island and Central America, Colombia, Peru, and Ecuador, including the Galapagos Islands (see Table 2, AMOVA Φst (485,1987) = 0.3268466, Exact Test P = 0.00001). We found that an IBD model does fit the structure of avian Haemoproteus genetic distances

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Journal Pre-proof (r=0.2082781, P = 0.003599, Perm= 10000). While avian Haemoproteus is differentiated from other Haemoproteus reported for the Galapagos Islands (Φst = 0.57, P = 0.00001), Haemoproteus found in birds of Gorgona Island shows a lower differentiation to mainland such parasites (Φst = 0.27-0.33, P = 0.00001).

3.5.

Genetic Variation among Avian-Groups

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There is low genetic differentiation of Haemoproteus found in birds within Gorgona Island (π =

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0.006111; Tajima´s D= -1.0808, P = 0.1419) and parasites of the same genera reported for

(183,1115) =0.0043514, Exact

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resident, migratory, marine and shorebirds species (see Appendix Table B5, AMOVA, Φst Test P = 0.00001). Haemoproteus haplogroups found on Gorgona

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Island are differentiated from those reported on migratory bird species (Φst = 0.57, P = 0.00001),

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marine and shorebirds (Φst = 0.53, P = 0.00001), but it is little differentiated from Haemoproteus

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4. Discussion

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(see Appendix, Table B5).

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in terrestrial bird species resident to mainland nearby Gorgona Island (Φst = 0.36, P = 0.00001)

This is the first study to examine the genetic variation of obligate blood parasites that may constitute potential pathogens of avian malaria to resident populations on Gorgona Island, Colombia. With the use of mtDNA amplicons, the genera Haemoproteus was identified in all PCR positive samples. Surprisingly, total prevalence on the island (57.9%) and the average prevalence by host species (42%) were higher than that the local average prevalence in continental areas (31%) (Belo et al., 2011; Gonzalez-Quevedo et al., 2016; González et al., 2015; Harrigan et al., 2014; Mantilla et al., 2013a, 2013b; Merino et al., 2000; Mijares et al., 2012) or

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Journal Pre-proof that seen in islands of the Lesser Antilles (28%) (Fallon et al., 2005). There is a host species specific effect in parasite prevalence in endemic birds of Gorgona Island; in particular, C. cyaneus gigas is the most prone to Haemoproteus infection among the island endemics. A similar host species effect in prevalence has been also documented in the Lesser Antilles (Fallon et al., 2003). The host species effect in prevalence further indicates a lower prevalence of avian

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populations in the Lesser Antilles (Fallon et al., 2003).

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Haemosporidia on C. flaveola in the island of Gorgona when compared with conspecific

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The differential prevalence between geographically distant populations may indicate variation in

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ecological conditions for obligate blood parasites. It has been previously suggested that habitat heterogeneity for avian Haemosporidia may gradually be conducive to divergence in

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immunological responses among hosts in islands as compared to conspecifics in the mainland

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(Beadell et al., 2007; Hart, 1990). The detection of only Haemoproteus in the samples could be due to the fact that it is easier to detect with molecular techniques (Clark et al., 2014), and if

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another genus, such as Plasmodium or mixed infections were present, its intensity circulating

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among island’s birds must have been very low in order to not be detected (Ciloglu et al., 2019; Harrigan et al., 2014).

It remains to be tested if non-resident birds are being a potential source of avian Haemoproteus, many migratory, marine and shorebirds remain to be examined for Haemosporidia in the island. The proximity of the island to mainland and the pattern of parasite genetic structure suggest that mainland is likely the main source of avian Haemoproteus. To us, it is unlikely that Haemoproteus in the Gorgona Island was colonized from outside the ETP by a long-distance 12

Journal Pre-proof transmitted parasite. In contrast, the avian fauna on Gorgona was likely colonized by haemosporidian parasites with both an amplitude of hosts and a wide range in Colombia and also in the ETP. Certainly, our results strongly suggest that Haemoproteus haplogroups H4 and H39 are generalists in terms of host amplitude and geographic breadth. Nevertheless, H502 Haemoproteus haplogroup is inexistent across the ETP but is found to Gorgona Island. Haemoproteus in this island H502 have over 0.7% sequence divergence to other haplogroups.

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Such a level of differentiation is within the higher bound of cyt b genetic distances (0.5-1%)

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among Haemoproteus species (Bensch et al., 2000; Reullier et al., 2006). The morphological

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description of H502 by microscopical techniques may prove valuable to support a potential novel

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Haemoproteus species or lineage (Mantilla et al., 2016).

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The reduced richness of flora and fauna in the Gorgona Island as compared to vast mainland extent (Giraldo and Garcés, 2012; Yockteng and Cavelier, 1998), strongly suggests that the 14

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resident birds on the island constitute a limited host opportunity to obligate blood parasites. Such

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a pattern may have resulted in accumulation of single nucleotide polymorphism following an IBD model, which may in turn explain the divergence of a possible novel haemosporidian species or lineage on Gorgona Island. We speculate that potential vectors belonging to the families Culicidae, Ceratopogonidae and Simuliidae reported on the island (Giraldo and Valencia, 2012; Longo and Blanco, 2014; “Plan de Manejo del Parque Nacional Natural Gorgona 2018-2023,” 2018) are also a limiting factor for transmission of Haemoproteus into the avian fauna of the Gorgona Island.

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Journal Pre-proof We found that avian Haemoproteus exhibit extraordinary high prevalence on the island and two of the haplotype groups found on this locality show high amplitude of hosts or large geographic breadth. The spatial genetic structure in the ETP also supports endemism of cyt b gene variation of avian Haemoproteus on Gorgona Island. For future studies, it would be important to implement the use of new molecular and microscopy techniques to determine whether or not haemosporidians constitute an actual threat to bird population on Gorgona Island. In conclusion,

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we provide evidence that these types of studies are important to show how insular avian

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communities are exposed to potential infectious diseases. Since avian malaria continue to

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constitute a poorly known, but a significant determinant for the extinction of island populations

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(García-Longoria et al., 2015), our methods and results can be instrumental to the conservation

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Acknowledgments

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strategies of populations of important bird areas in the ETP.

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We are grateful to field assistants N. Pérez, L. Calvert, H. Calero, and C. Guerrero. We are indebted to M. Mosquera-Escudero and the facilities of the Department of Physiological Sciences, Univalle, and we thank C. Muñoz for her labwork assistance. We thank L. Payan and X. Zorrilla kind administrative personnel of the Gorgona Island National Park. Collected specimens were deposited in the Colección Ornitológica Univalle.

Funding: The research was supported by Universidad del Valle, Cali, Colombia.

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Journal Pre-proof Tables Table 1. Information about prevalence. Prevalence of avian Haemoproteus on Gorgona Island, Colombia.

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Infected

Prevalence (%)

Actitis macularius Amazilia tzacatl Thamnophilus atrinucha gorgonae Tyrannus melancholicus Coereba flaveola gorgonae Cyanerpes cyaneus gigas Sporophila luctuosa

5 2 11 1 11 25 1

1 0 3 1 4 22 1

20,0 27,3 -b 36,4 88,0 -b

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2 1 2 13

2 1

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A short positive Haemoproteus sequence from an individual of Catharus ustulatus was excluded from analyses

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Specific prevalence was not calculated for sample sizes <5

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b

Haplogroups Gorgona Island 39 4 502 1

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Species a

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Journal Pre-proof Table 2. AMOVA results. Descriptive statistics for the cyt b gene alignment of 78 Haemoproteus haplogroups in the eastern tropical Pacific and regions of the world. Variable sites of cyt b gene (s), the richness of haplotype groups (h), expected heterozygosity (He), nucleotide diversity (π) and Tajima's D test. Areas

s

h

π (+/- s.d.)

He

Tajima´s D

p-value

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Comparison within the Eastern Tropical Pacific (ETP) 77

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0.05597

0.056798 +/- 0.029109

0.9056

0.8587

Colombia-Ecuador-Peru

158

69

0.04345

0.043350 +/-0.021336

-0.96244

0.165

Gorgona Island

45

3

0.01232

0.012320 +/-0.006821

-2.04607

0.0076

Galápagos Islands Regional Comparison Worldwide

76

9

0.07974

1.90393

0.9877

ETP

163

77

0.05570

0.056914 +/- 0.027760

-0.29598

0.47000

Africa

75

10

0.05168

0.049199 +/- 0.026870

-0.52528

0.29800

Asia

67

10

0.05437

0.054326 +/- 0.029505

-0.18195

0.45800

Europe

75

Oceania

64

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Mexico-Panama-Costa Rica

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0.079133 +/-0.040280

0.05226

0.053375 +/- 0.028036

0.13200

0.62700

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0.05542

0.049082 +/- 0.027162

-0.40350

0.34400

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References Alley, M.R., Fairley, R.A., Martin, D.G., Howe, L., Atkinson, T., 2008. An outbreak of avian malaria in captive yellowheads/mohua (mohoua ochrocephala). N. Z. Vet. J. 56, 247–251. https://doi.org/10.1080/00480169.2008.36842 Apanius, V., Yorinks, N., Bermingham, E., Ricklefs, R.E., 2000. Island and taxon effects in parasitism and resistance of Lesser Antillean birds. Ecology 81, 1959–1969.

of

https://doi.org/10.1890/0012-9658(2000)081[1959:IATEIP]2.0.CO;2

ro

Atkinson, C.T., Dusek, R.J., Woods, K.L., Iko, W.M., 2000. Pathogenicity of Avian Malaria in

re

https://doi.org/10.7589/0090-3558-36.2.197

-p

Experimentally-Infected Hawaii Amakihi. J. Wildl. Dis. 36, 197–201.

lP

Beadell, J.S., Atkins, C., Cashion, E., Jonker, M., Fleischer, R.C., 2007. Immunological change in a parasite-impoverished environment: Divergent signals from four island taxa. PLoS One

na

2. https://doi.org/10.1371/journal.pone.0000896 Beadell, J.S., Ishtiaq, F., Covas, R., Melo, M., Warren, B.H., Atkinson, C.T., Bensch, S., Graves,

ur

G.R., Jhala, Y. V, Peirce, M.A., Rahmani, A.R., Fonseca, D.M., Fleischer, R.C., 2006.

Jo

Global phylogeographic limits of Hawaii’s avian malaria. Proc. R. Soc. B Biol. Sci. 273, 2935–2944. https://doi.org/10.1098/rspb.2006.3671 Belo, N.O., Pinheiro, R.T., Reis, E.S., Ricklefs, R.E., Braga, É.M., 2011. Prevalence and lineage diversity of avian haemosporidians from three distinct cerrado habitats in Brazil. PLoS One 6, 1–8. https://doi.org/10.1371/journal.pone.0017654 Bensch, S., Hellgren, O., Pérez-Tris, J., 2009. MalAvi: A public database of malaria parasites and related haemosporidians in avian hosts based on mitochondrial cytochrome b lineages. Mol. Ecol. Resour. 9, 1353–1358. https://doi.org/10.1111/j.1755-0998.2009.02692.x

17

Journal Pre-proof Bensch, S., Stjernman, M., Hasselquist, D., Ostman, O., Hansson, B., Westerdahl, H., Pinheiro, R.T., 2000. Host specificity in avian blood parasites : a study of Plasmodium and Haemoproteus mitochondrial DNA amplified from birds. Proc. R. Soc. 267, 1583-1589. https://doi.org/10.1098/rspb.2000.1181 Benson, D.A., Karsch-Mizrachi, I., Lipman, D.J., Ostell, J., Sayers, E.W., 2011. GenBank. Nucleic Acids Res. 39, D32–D37. https://doi.org/10.1093/nar/gkq1079

of

Blanco, J.F., 2009. The hydroclimatology of Gorgona Island: Seasonal and ENSO-related

ro

patterns. Actual. Biológicas 31, 111–121.

-p

Bohonak, A.J., 2002. IBD (Isolation by Distance): A Program for Analyses of Isolation by

re

Distance. J. Hered. 93, 153–154. https://doi.org/10.1093/jhered/93.2.153 Cadena Ortiz, H.F., 2015. Malaria Aviar en los Chingolos (Zonotrichia capensis) del Parque

lP

Recreacional-Bosque Protector Jerusalem, Pichincha, Ecuador.

na

Chasar, A., Loiseau, C., Valkiunas, G., Iezhova, T., Smith, T.B., Sehgal, R.N.M., 2009. Prevalence and diversity patterns of avian blood parasites in degraded African rainforest

ur

habitats. Mol. Ecol. 18, 4121–4133. https://doi.org/10.1111/j.1365-294X.2009.04346.x

Jo

Ciloglu, A., Ellis, V.A., Bernotienė, R., Valkiūnas, G., Bensch, S., Waldenström, J., Bensch, S., Hasselquist, D., Östman, Ö., 2019. A New Nested Polymerase Chain Reaction Method Very Efficient in Detecting Plasmodium and Haemoproteus Infections From Avian Blood. Parasitol. Res. 118, 191–201. https://doi.org/10.1645/ge-3221rn Clark, N.J., Clegg, S.M., Lima, M.R., 2014. A review of global diversity in avian haemosporidians (Plasmodium and Haemoproteus: Haemosporida): New insights from molecular data. Int. J. Parasitol. 44, 329–338. https://doi.org/10.1016/j.ijpara.2014.01.004 Drummond, A.J., Suchard, M.A., Xie, D., Rambaut, A., 2012. Bayesian phylogenetics with

18

Journal Pre-proof BEAUti and the BEAST 1.7. Mol. Biol. Evol. 29, 1969–1973. https://doi.org/10.1093/molbev/mss075 Edgar, R.C., 2010. Search and clustering orders of magnitude faster than BLAST. Bioinformatics 26, 2460–2461. https://doi.org/10.1093/bioinformatics/btq461 Ewen, J.G., Bensch, S., Blackburn, T.M., Bonneaud, C., Brown, R., Cassey, P., Clarke, R.H., Pérez-Tris, J., 2012. Establishment of exotic parasites: The origins and characteristics of an

of

avian malaria community in an isolated island avifauna. Ecol. Lett. 15, 1112–1119.

ro

https://doi.org/10.1111/j.1461-0248.2012.01833.x

-p

Excoffier, L., Lischer, H.E.L., 2010. Arlequin suite ver 3.5: A new series of programs to perform

re

population genetics analyses under Linux and Windows. Mol. Ecol. Resour. 10, 564–567. https://doi.org/10.1111/j.1755-0998.2010.02847.x

lP

Fallon, S.M., Bermingham, E., Ricklefs, R.E., 2005. Host specialization and geographic

na

localization of avian malaria parasites: a regional analysis in the Lesser Antilles. Am. Nat. 165, 466–480. https://doi.org/10.1086/428430

ur

Fallon, S.M., Bermingham, E., Ricklefs, R.E., 2003. Island and taxon effects in parasitism

Jo

revisited: avian malaria in the Lesser Antilles. Evolution 57, 606–615. https://doi.org/10.1554/0014-3820(2003)057 García-Longoria, L., Magallanes, S., de Lope, F., Marzal, A., 2015. Biological invasions of malaria parasites and their bird hosts, Biological Invasions: Patterns, Management and Economic Impacts. https://doi.org/10.1108/09675420910984664 Gil-Vargas, D.L., Sedano-Cruz, R.E., 2019. Genetic variation of avian malaria in the tropical Andes: a relationship with the spatial distribution of hosts. Malar. J. 18, 129. https://doi.org/10.1186/s12936-019-2699-9

19

Journal Pre-proof Giraldo, A., Diazgranados, M.C., Gutiérez-Landázuri, C., 2014. Isla Gorgona, enclave estratégico para los esfuerzos de conservación en el Pacífico Oriental Tropical. Rev. Biol. Trop. 62, 1–12. Giraldo, A., Garcés, M., 2012. Vertebrados del ambiente terrestre del Parque Nacional Natural Gorgona, in: Isla Gorgona, Paraíso de Biodiversidad y Ciencia. Giraldo, A., Valencia, B., 2012. Artropofauna del PNN Gorgona, in: Valle, P.E.U. del (Ed.), Isla

of

Gorgona, Paraíso de Biodiversidad y Ciencia. Cali, p. 221.

ro

Gonzalez-Quevedo, C., Rivera-Gutierrez, H.F., Pabón, A., 2016. Prevalence of haemosporidians

-p

in a Neotropical endemic bird area. Avian Conserv. Ecol. 11. https://doi.org/10.5751/ACE-

re

00834-110107

González, A.D., Lotta, I.A., García, L.F., Moncada, L.I., Matta, N.E., 2015. Avian

lP

haemosporidians from Neotropical highlands: Evidence from morphological and molecular

na

data. Parasitol. Int. 64, 48–59. https://doi.org/10.1016/j.parint.2015.01.007 Harrigan, R.J., Sedano, R., Chasar, A.C., Chaves, J. a., Nguyen, J.T., Whitaker, A., Smith, T.B.,

ur

2014. New host and lineage diversity of avian haemosporidia in the northern Andes. Evol.

Jo

Appl. 1–13. https://doi.org/10.1111/eva.12176 Hart, B.L., 1990. Behavioral adaptations to pathogens and parasites: Five strategies. Neurosci. Biobehav. Rev. 14, 273–294. https://doi.org/10.1016/S0149-7634(05)80038-7 Hellgren, O., Waldenström, J., Bensch, S., 2004. A NEW PCR ASSAY FOR SIMULTANEOUS STUDIES OF LEUCOCYTOZOON, PLASMODIUM, AND HAEMOPROTEUS FROM AVIAN BLOOD. J. Parasitol. 90, 797–802. https://doi.org/10.1645/ge-184r1 Jones, M.R., Cheviron, Z. a, Carling, M.D., 2013. Spatial patterns of avian malaria prevalence in Zonotrichia capensis on the western slope of the Peruvian Andes. J. Parasitol. 99, 903–5.

20

Journal Pre-proof https://doi.org/10.1645/12-147.1 Kamiya, T., O’Dwyer, K., Nakagawa, S., Poulin, R., Clark, N.J., 2014. Host diversity drives parasite diversity: Meta-analytical insights into patterns and causal mechanisms. Ecography (Cop.). 37, 689–697. https://doi.org/10.1111/j.1600-0587.2013.00571.x Letunic, I., Bork, P., 2007. Interactive Tree Of Life (iTOL): An online tool for phylogenetic tree display and annotation. Bioinformatics 23, 127–128.

of

https://doi.org/10.1093/bioinformatics/btl529

ro

Levin, I.I., Outlaw, D.C., Vargas, F.H., Parker, P.G., 2009. Plasmodium blood parasite found in

re

https://doi.org/10.1016/j.biocon.2009.06.017

-p

endangered Galapagos penguins (Spheniscus mendiculus). Biol. Conserv. 142, 3191–3195.

Levin, I.I., Zwiers, P., Deem, S.L., Geest, E. a., Higashiguchi, J.M., Iezhova, T. a., Jiménez-

lP

Uzcátegui, G., Kim, D.H., Morton, J.P., Perlut, N.G., Renfrew, R.B., Sari, E.H.R.,

na

Valkiunas, G., Parker, P.G., 2013. Multiple Lineages of Avian Malaria Parasites (Plasmodium) in the Galapagos Islands and Evidence for Arrival via Migratory Birds.

ur

Conserv. Biol. 27, 1366–1377. https://doi.org/10.1111/cobi.12127

Jo

Levine, N.D., 2018. The Protozoan Phylum Apicomplexa, The Protozoan Phylum Apicomplexa Volume II. CRC Press. https://doi.org/10.1201/9781351076111 Longmire, J.L., Gee, G.F., Hardekopf, C.L., Graham, A., Mark, G.A., 1992. Establishing Paternity in Whooping Cranes ( Grus americana ) by DNA Analysis Published by : American Ornithological Society Stable URL : http://www.jstor.org/stable/4088366 REFERENCES Linked references are available on JSTOR for this article : You may nee. Auk 109, 522–529. Longo, M., Blanco, J.F., 2014. Discourseontology, the subject principle, perspectivation: An

21

Journal Pre-proof integrative agent-centered sociocognitive perspective on translating. Rev. Biol. Trop. 62, 85–105. https://doi.org/10.20916/1812-3228-2017-4-69-79 Lotta, I.A., Valkiūnas, G., Pacheco, M.A., Escalante, A.A., Hernández, S.R., Matta, N.E., 2019. Disentangling Leucocytozoon parasite diversity in the neotropics: Descriptions of two new species and shortcomings of molecular diagnostics for leucocytozoids. Int. J. Parasitol. Parasites Wildl. 9, 159–173. https://doi.org/10.1016/j.ijppaw.2019.05.002

of

Mantilla, J.S., González, A.D., Lotta, I.A., Moens, M., Pacheco, M.A., Escalante, A.A.,

ro

Valkiunas, G., Moncada, L.I., Pérez-Tris, J., Matta, N.E., 2016. Haemoproteus

-p

erythrogravidus n. sp. (Haemosporida, Haemoproteidae): Description and molecular

re

characterization of a widespread blood parasite of birds in South America. Acta Trop. 159, 83–94. https://doi.org/10.1016/j.actatropica.2016.02.025

lP

Mantilla, J.S., González, A.D., Valki, G., 2013a. Description and molecular characterization of

na

Plasmodium ( Novyella ) unalis sp . nov . from the Great Thrush ( Turdus fuscater ) in highland of Colombia. Parasitol. Res. Volume 112, 4193–4204.

ur

https://doi.org/10.1007/s00436-013-3611-0

Jo

Mantilla, J.S., Matta, N.E., Pacheco, M.A., Escalante, A.A., González, A.D., Moncada, L.I., 2013b. Identification of Plasmodium (Haemamoeba) lutzi (Lucena, 1939) from Turdus fuscater (Great Thrush) in Colombia. J. Parasitol. 99, 662–668. https://doi.org/10.1645/12138.1 Merino, S., Moreno, J., Sanz, J., Arriero, E., 2000. Are avian blood parasites pathogenic in the wild? A medication experiment in blue tits (Parus caeruleus). R. Soc. 94, 32–41. https://doi.org/10.1098/rspb.2000.1312 Mijares, A., Rosales, R., Silva-Iturriza, A., 2012. Hemosporidian Parasites in Forest Birds from

22

Journal Pre-proof Venezuela: Genetic Lineage Analyses. Avian Dis. Dig. 7, e56–e57. https://doi.org/10.1637/10223-1005812-DIGEST.1 Miller, M.A., Pfeiffer, W., Schwartz, T., 2010. Creating the CIPRES Science Gateway for inference of large phylogenetic trees, in: Gateway Computing Environments Workshop. https://doi.org/10.1109/GCE.2010.5676129 Niebuhr, C.N., Poulin, R., Tompkins, D.M., 2016. Is avian malaria playing a role in native bird

ro

1–15. https://doi.org/10.1371/journal.pone.0165918

of

declines in New Zealand? Testing hypotheses along an elevational gradient. PLoS One 11,

-p

Plan de Manejo del Parque Nacional Natural Gorgona 2018-2023, 2018.

re

Rangel-Ch., J.O., 1995. ISLAS DE GORGONA Y GORGONILLA, in: Colombia Diversidad Biotica I. pp. 145–154. https://doi.org/10.1117/12.2055279

lP

Reullier, J., Pérez-Tris, J., Bensch, S., Secondi, J., 2006. Diversity , distribution and exchange of

na

blood parasites meeting at an avian moving contact zone 15, 753–763. https://doi.org/10.1111/j.1365-294X.2005.02826.x

ur

Ricklefs, R.E., Outlaw, D.C., 2010. A molecular clock for malaria parasites. Science 329, 226–

Jo

229. https://doi.org/10.1126/science.1188954 Sedano, R.E., Burns, K.J., 2010. Are the Northern Andes a species pump for Neotropical birds? Phylogenetics and biogeography of a clade of Neotropical tanagers (Aves: Thraupini). J. Biogeogr. 37, 325–343. https://doi.org/10.1111/j.1365-2699.2009.02200.x Swofford, D.L., 2002. Phylogenetic Analysis Using Parsimony. Options 42, 294–307. https://doi.org/10.1007/BF02198856 Tajima, F., 1989. Statistical method for testing the neutral mutation hypothesis by DNA polymorphism. Genetics 123, 585–595. https://doi.org/PMC1203831

23

Journal Pre-proof Tamura, K., Stecher, G., Peterson, D., Filipski, A., Kumar, S., 2013. MEGA6: Molecular Evolutionary Genetics Analysis Version 6.0. Mol. Biol. Evol. 30, 2725–2729. https://doi.org/10.1093/molbev/mst197 Tavaré, S., 1986. Some Probabilistic and Statistical Problems in the Analysis of DNA Sequences, Some Mathematical Questions in Biology--DNA Sequence Analysis. Thayer, J.E., Bangs, O., 1905. The Vertebrata of Gorgona Island, Colombia, in: Bulletin of the

of

Museum At Harvard College. pp. 46: 91-98.

ro

Tompkins, D.M., Gleeson, D.M., 2006. Relationship between avian malaria distribution and an

-p

exotic invasive mosquito in New Zealand. J. R. Soc. New Zeal. 36, 51–62. https://doi.org/10.1080/03014223.2006.9517799

re

Valkiūnas, G., 2005. Avian malaria parasites and other Haemosporidia, CRC Press.

lP

https://doi.org/10.1201/9780203643792.fmatt

na

Waldenström, J., Bensch, S., Hasselquist, D., Östman, Ö., 2004. A New Nested Polymerase Chain Reaction Method Very Efficient in Detecting Plasmodium and Haemoproteus

ur

Infections From Avian Blood. J. Parasitol. 90, 191–194. https://doi.org/10.1645/GE-

Jo

3221RN

Yockteng, R., Cavelier, J., 1998. Diversidad y mecanismos de dispersión de árboles de la Isla Gorgona y de los bosques húmedos tropicales del Pacífico colombo-ecuatoriano. Rev. Biol. Trop. 46, 45–53.

24

Journal Pre-proof List of Figures

Figure 1. Map of avian haemosporidia known distribution across the eastern tropical Pacific region (red dots). Distribution of Haemoproteus mtDNA sequences from GenBank and MalAvi databases including those found in Gorgona Island, and further adjacent areas to the ETP

of

(grey dots).

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Figure 2. Genealogy of 78 avian Haemoproteus cyt b gene haplogroups of the eastern

-p

tropical Pacific region. The Bayesian inference consensus tree shows ≥0.95 posterior

re

probability support to nodes. Haplogroups are shown at the tips of the tree (see Table S2 in supplementary material), but also the GenBank accession number, a short identification of the

lP

known Haemosporidia genera o subgenera, and a two-letter country code for BE (Belize), CO

na

(Colombia), CR (Costa Rica), EC (Ecuador), GAL (Galapagos Island-Ecuador), MX (Mexico), PA (Panama), and PE (Peru). Divergence time is shown for each of the two clades containing the

Jo

ur

three haplogroups (bold font) found in Gorgona island.

25

Journal Pre-proof Tabla B1. Haemoproteus sequence used to perform AMOVA by continent

-p

ro

of

Reference (Križanauskienė et al., 2006) (Križanauskienė et al., 2006) (Križanauskienė et al., 2006) (Ishtiaq et al., 2018) (Ishtiaq et al., 2018) (Nourani et al., 2018) (Nourani et al., 2018) (Nourani et al., 2018) (Nourani et al., 2018) (Nourani et al., 2018) (Dimitrov et al., 2013) (Dimitrov et al., 2013) (Hellgren et al., 2007) (Hellgren et al., 2007) (Hellgren et al., 2007) (Hellgren et al., 2007) (Hellgren et al., 2007) (Hellgren et al., 2007) (Hellgren et al., 2007) (Dimitrov et al., 2013) (Dimitrov et al., 2013) (Hellgren et al., 2007) (Dimitrov et al., 2013) (Loiseau et al., 2017) (Loiseau et al., 2017) (Loiseau et al., 2017) (Musa et al., 2019) (Musa et al., 2019) (Musa et al., 2019) (Chaisi et al., 2019) (Chaisi et al., 2019) (Chaisi et al., 2019) (Chaisi et al., 2019) (Fecchio et al., 2019) (Fecchio et al., 2019) (Fecchio et al., 2019) (Fecchio et al., 2019) (Fecchio et al., 2019) (Fecchio et al., 2019)

lP

re

Continent Asia Asia Asia Asia Asia Asia Asia Asia Asia Asia Europe Europe Europe Europe Europe Europe Europe Europe Europe Europe Europe Europe Europe Africa Africa Africa Africa Africa Africa Africa Africa Africa Africa Oceania Oceania Oceania Oceania Oceania Oceania

ur

na

Parasite Genus Haemoproteus Haemoproteus Haemoproteus Haemoproteus Haemoproteus Haemoproteus Haemoproteus Haemoproteus Haemoproteus Haemoproteus Haemoproteus Haemoproteus Haemoproteus Haemoproteus Haemoproteus Haemoproteus Haemoproteus Haemoproteus Haemoproteus Haemoproteus Haemoproteus Haemoproteus Haemoproteus Haemoproteus Haemoproteus Haemoproteus Haemoproteus Haemoproteus Haemoproteus Haemoproteus Haemoproteus Haemoproteus Haemoproteus Haemoproteus Haemoproteus Haemoproteus Haemoproteus Haemoproteus Haemoproteus

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GenBank Acession DQ368341 DQ368343 DQ368357 KY623720 KY623721 MG976511 MG976519 MG976526 MG976536 MG976558 AF495579 AF495580 AY831760.1 DQ000320 DQ000322 DQ058614 DQ368347 DQ368348 DQ368360 DQ368371 DQ368372 DQ847203 KC568475 KT376897 KT376908 KT376927 MF442567 MF442573 MF442583 MH492267 MH492269 MH492272 MH492291 AY714135 AY714174 AY714179 JX021537 JX021550 KU296186

26

Journal Pre-proof KX100323 MG387220 MG387222

Haemoproteus Haemoproteus Haemoproteus

(Fecchio et al., 2019) (Fecchio et al., 2019) (Fecchio et al., 2019)

Oceania Oceania Oceania

Jo

ur

na

lP

re

-p

ro

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References Chaisi, M.E., Osinubi, S.T., Dalton, D.L., Suleman, E., 2019. Occurrence and diversity of avian haemosporidia in Afrotropical landbirds. Int. J. Parasitol. Parasites Wildl. 8, 36–44. https://doi.org/10.1016/j.ijppaw.2018.12.002 Dimitrov, D., Valkiunas, G., Zehtindjiev, P., Ilieva, M., Bensch, S., 2013. Molecular characterization of haemosporidian parasites (Haemosporida) in yellow wagtail (Motacilla flava), with description of in vitro ookinetes of Haemoproteus motacillae. Zootaxa 3666, 369–381. https://doi.org/10.11646/zootaxa.3666.3.7 Fecchio, A., Wells, K., Bell, J.A., Tkach, V. V., Lutz, H.L., Weckstein, J.D., Clegg, S.M., Clark, N.J., 2019. Climate variation influences host specificity in avian malaria parasites. Ecol. Lett. 22, 547–557. https://doi.org/10.1111/ele.13215 Hellgren, O., Waldenström, J., Peréz-Tris, J., Szöll Ösi, E., Hasselquist, D., Krizanauskiene, A., Ottosson, U., Bensch, S., 2007. Detecting shifts of transmission areas in avian blood parasites - A phylogenetic approach. Mol. Ecol. 16, 1281–1290. https://doi.org/10.1111/j.1365-294X.2007.03227.x Ishtiaq, F., Rao, M., Palinauskas, V., 2018. Molecular characterization and morphological description of cryptic haemoproteids in the laughingthrushes (Leiothrichidae) in the western and eastern Himalaya, India [version 1; referees: 3 approved]. Wellcome Open Res. 3, 1– 16. https://doi.org/10.12688/wellcomeopenres.14675.1 Križanauskienė, A., Hellgren, O., Kosarev, V., Sokolov, L., Bensch, S., Valkiūnas, G., 2006. Variation in Host Specificity Between Species of Avian Hemosporidian Parasites: Evidence From Parasite Morphology and Cytochrome B Gene Sequences . J. Parasitol. 92, 1319– 1324. https://doi.org/10.1645/ge-873r.1 Loiseau, C., Melo, M., Lobato, E., Beadell, J.S., Fleischer, R.C., Reis, S., Doutrelant, C., Covas, R., 2017. Insularity effects on the assemblage of the blood parasite community of the birds from the Gulf of Guinea. J. Biogeogr. 44, 2607–2617. https://doi.org/10.1111/jbi.13060 Musa, S., Mackenstedt, U., Woog, F., Dinkel, A., 2019. Avian malaria on Madagascar: prevalence, biodiversity and specialization of haemosporidian parasites. Int. J. Parasitol. 49, 199–210. https://doi.org/10.1016/j.ijpara.2018.11.001 Nourani, L., Aliabadian, M., Mirshamsi, O., Djadid, N.D., 2018. Molecular detection and genetic diversity of avian haemosporidian parasites in iran. PLoS One 13, 1–16. https://doi.org/10.1371/journal.pone.0206638

27

Journal Pre-proof Table B2. Individuals examined in Gorgona Island. Sample identification by accession number of the GenBank, field number, catalog of CAUV for collected specimens, the number of PCR assays per sample and overall PCR result for Haemosporidia. Field Number

PCR Assays

Haemosporidia

-

A207

3

-

Q416

3

-

Q430

3

-

Actitis macularius

MF990734

Q438

2

+

Actitis macularius

-

Q440

3

-

NPA 021

1

-

3

-

3

+

2

+

2

-

A179

2

-

A185

3

-

A189

2

-

A39

2

+

1

-

Actitis macularius Actitis macularius

Amazilia tzacalt Amazilia tzacalt

Coereba flaveola gorgonae

Sequence no submitted to the GenBank MF990714

Q410

-

A122

Coereba flaveola gorgonae Coereba flaveola gorgonae Coereba flaveola gorgonae Coereba flaveola gorgonae

A110

-p

Catharus ustulatus

Q443

ro

Actitis macularius

Catalog CAUV

of

GenBank Accession

re

Spe cies

MF990721

Coereba flaveola gorgonae

-

Coereba flaveola gorgonae

Sequence not submitted to the GenBank

Q405

3

+

-

Q406

2

-

Coereba flaveola gorgonae

MF990733

Q421

1

+

Coereba flaveola gorgonae

-

Q424

3

-

Sequence not submitted to the GenBank

NPA 017

1

+

-

NPA 019

1

-

NPA 020

1

+

NPA 058

2

+

Cyanerpes cyaneus gigas

Sequence not submitted to the GenBank Sequence not submitted to the GenBank MF990712

A102

2

+

Cyanerpes cyaneus gigas

MF990713

A106

3

+

Cyanerpes cyaneus gigas

MF990715

A111

3

+

Cyanerpes cyaneus gigas

MF990716

A115

3

+

Cyanerpes cyaneus gigas

MF990719

A176

2

+

Cyanerpes cyaneus gigas

MF990720

A229

2

+

Cyanerpes cyaneus gigas

-

A242

2

-

Cyanerpes cyaneus gigas

MF990722

A52

2

+

Cyanerpes cyaneus gigas Cyanerpes cyaneus gigas Cyanerpes cyaneus gigas

na

ur

Cyanerpes cyaneus gigas

Jo

Coereba flaveola gorgonae

lP

Coereba flaveola gorgonae

NPA 056

7339-06119

28

Field Number

Cyanerpes cyaneus gigas

MF990723

Cyanerpes cyaneus gigas Cyanerpes cyaneus gigas

Haemosporidia

A55

2

+

MF990725

A86

2

+

A91

2

+

A93

2

+

Cyanerpes cyaneus gigas

Sequence not submitted to the GenBank Sequence not submitted to the GenBank MF990726

A99

1

+

Cyanerpes cyaneus gigas

MF990728

Q398

2

+

Cyanerpes cyaneus gigas

Sequence not submitted to the GenBank MF990731

Q401

2

+

Q409

2

+

Sequence not submitted to the GenBank MF990732

Q412

2

+

2

+

Cyanerpes cyaneus gigas

-

Q426

2

-

Cyanerpes cyaneus gigas

Sequence not submitted to the GenBank Sequence not submitted to the GenBank MF990727

Q427

2

+

Q434

2

+

Q397

2

+

A121

2

-

A143

2

+

-

A35

2

+

-

A73

2

-

MF990724

A81

2

+

-

NPA 054

7334-06116

2

-

-

NPA 055

7337-06117

2

-

-

NPA 057

7333-06113

2

-

-

NPA 059

7338-06118

2

-

-

NPA 060

7334-06114

2

-

-

A0073

2

-

MF990729

Q399

2

+

Cyanerpes cyaneus gigas Sporophila luctuosa Thamnophilus atrinucha gorgonae Thamnophilus atrinucha gorgonae Thamnophilus atrinucha gorgonae Thamnophilus atrinucha gorgonae Thamnophilus atrinucha gorgonae Thamnophilus atrinucha gorgonae Thamnophilus atrinucha gorgonae Thamnophilus atrinucha gorgonae Thamnophilus atrinucha gorgonae Thamnophilus atrinucha gorgonae Thamnophilus atrinucha gorgonae Tyrannus melancholicus

MF990718

Q419

-p

Cyanerpes cyaneus gigas

lP

Cyanerpes cyaneus gigas

na

Cyanerpes cyaneus gigas

ur

Cyanerpes cyaneus gigas

Catalog CAUV

ro

GenBank Accession

Jo

Spe cies

of

PCR Assays

re

Journal Pre-proof

29

Journal Pre-proof Table B3. Haplotype group determination using USEARCH. Country and accession number in GenBank (* centroids for each haplotype group). Haplogroups

Country Antilles Brazil

of

Colombia

GenBank Accession GQ141565 HQ287537 JX029911 KF537317 MF990721 MF990730 KJ661305 KJ661311 KJ661316 KJ661317 KT373859 MF077652 MF077671 MF077676 DQ241548 DQ241549

ro

Ecuador

-p

4

re

United States

na

lP

French Guiana Norte America Continental

Jo

ur

Uruguay Antilles

5

Ecuador United States

6

Colombia Antilles

7

Colombia

Ecuador

GU252004* DQ241545 DQ241547 AY167243 GQ141568 GU251995* GQ395638 GQ395652 GQ141566 KF537319 KF537321* AY455658 KF537309 KF537318 KF537329* KF537330 KT698209 EF153649 30

Journal Pre-proof Country

GenBank Accession KJ661246 KJ661247 KJ661251 KJ661253 KJ661254 KJ661256 KJ661257 KJ661258 KJ661262 KJ661263 KJ661264 KJ661269* KJ661270* KJ661272 KJ661273 KJ661274 KJ661275 KJ661277 KJ661278 KJ661279 KJ661280 KJ661281 KJ661282 KJ661284 KJ661286 KJ661287 KJ661288 KJ661289 KJ661291 KJ661292 KJ661293 KJ661294 KJ661295 KJ661296 KJ661297 KJ661298 KJ661302 KJ661303

Jo

ur

na

lP

re

-p

ro

of

Haplogroups

31

Journal Pre-proof Country

GenBank Accession KJ661304 KJ661307 KJ661309 KJ661312 KJ661313 KJ661314 KJ661318 KJ661324 KJ661325 KU364580 KU364581 KU364582 KU364583 KU364584 EF153649 JQ764618 JQ988106 KF767417 JN819378* JN819383 JN819399 JQ988577 EF153648 EF153652 KC480265* KC480265* MF990712 MF990713 MF990714 MF990715 MF990716 MF990717 MF990718 MF990719 MF990720 MF990722 MF990723 MF990724

lP

re

Peru

-p

ro

of

Haplogroups

Costa Rica

na

14

ur

Peru

Jo

33

39

Ecuador Peru

Colombia

32

Journal Pre-proof

40

Colombia Ecuador Peru Costa Rica

42

-p

Ecuador Colombia

lP

re

45

Colombia

Jo

ur

na

46

Brazil

49

Colombia Peru Belize

56

GenBank Accession MF990725 MF990726 MF990728 MF990729 MF990731 MF990732* MF990733 MF990734 KM211348 KC480266* KC480266* JN819388* JN819393 KJ661259 KM211346* KM211349 KM211352 KF537300 KF537301 KF537302 KF537303 KF537306* KF537311 KU562167 KU562168 KU562169 KU562170 KF537320* KF537331 JQ988135 JF833043 JF833043 JF833044 JF833045 JF833048 JF833049* JF833050 JF833043

of

Country

ro

Haplogroups

Ecuador

United States

33

Journal Pre-proof Haplogroups

Country Panamá Colombia Costa Rica

Ecuador

61

ro

of

Peru

GenBank Accession JF833043 JF833058 KM211350 JN819345 JN819385 KT373861* KU364585 KU364586 JQ988150 JQ988167 JQ988254 JQ988745 JX029915 KC121053 KF537304 KX130087 KJ661301 KT373863 KU364540 KU364541 KU364542 KU364543 KU364544 KU364545 KU364546 KU364547 KU364548 KU364549 KU364550 KU364551 KU364552 KU364553 KU364554 KU364555 KU364556 KU364557 KU364558 KU364559

-p

Brazil

ur

na

lP

re

Colombia

Jo

65

Ecuador

34

Journal Pre-proof Country

GenBank Accession KU364560 KU364561 KU364562 KU364563 KU364564 KU364565 KU364566 KU364567 KU364568 KU364569 KU364570 KU364571 KU364572 KU364573 KU364574 KU364575 JQ988105 JQ988147 JQ988371 JQ988384 JQ988406 JQ988426 JQ988430 JQ988447 JQ988487 JQ988488 JQ988492* JQ988521 JQ988538 JQ988563 JQ988570 JQ988744 KF767420 JQ988144 JQ988323 JQ988370 JQ988393* JX029900

Peru

Jo

ur

na

lP

re

-p

ro

of

Haplogroups

66

Peru

69

Brazil

35

Journal Pre-proof Country

GenBank Accession JX029903 JX029920 KU562174 KU562175 KU562176 KU562177 KU562178 KU562183 KU562184 KU562185 KU562186 KU562187 JX029900 JQ988656* JQ988575* KU562241 KJ661283 JQ988571* DQ241534 DQ241553 DQ241554 KF767425* JQ988107 JQ988134* JQ988136 JQ988117* JQ988489 JQ988123* JQ988256* MF077663 KF767419* JX029910* JQ988585 KU562226 KU562227 KU562228 KU562229 KU562230

-p

Costa Rica Peru Peru Brazil Ecuador Peru

re

73

lP

74

Guyana

na

75

Peru

Jo

ur

77

Peru

78

Peru

79 80

Peru Peru United States Peru Brazil Peru

81 87

155

ro

of

Haplogroups

Brazil

36

Journal Pre-proof Haplogroups

Country French Guiana Peru Peru Peru

156 157

GenBank Accession DQ241558 KU562246* KU562248* KU562247* KU562129 KU562130 KU562131 KU562207 KT373865

Brazil 158

Peru Peru Brazil

lP

183

Colombia

Jo

ur

185 186

na

Ecuador

184

Ecuador

Colombia Ecuador Antilles Brazil

206 Costa Rica Norte America Continental 207 229

KU562244* KJ661326 KJ661328* JQ988222* KF767416* KU562136 KF537296 KF537299* KT373862 JF833060 JF833061* KF537325* JF833065* AF465579 AY167242 GU256263* HQ287540 JN819369 JN819373

ro

181 182

-p

Ecuador

DQ241556

re

178

of

Ecuador French Guiana Peru

Colombia Costa Rica Brazil

GQ141606 KF537282 KF537285 KF537295* JN819370 KU131583 37

Journal Pre-proof Country

243

Ecuador

246 249 250

Costa Rica Peru Peru Colombia United States

234

re

-p

237

of

241

Colombia Colombia Costa Rica Ecuador Peru Peru

lP

254

Peru

Jo

339 345 350 361 367

Peru Colombia Peru Ecuador Antilles Ecuador United States Colombia Costa Rica Mexico Colombia United States

na

ur

255 320 324 337

GenBank Accession KU131585* KJ644778 KF537326* JN819389* KJ661310 JN819389* KF482344* JF833051* JF833054 JF833055 JF833056 JF833059 JF833066 JN792143* JQ988462* JQ988446* KX130086 MF077670 KF767421 KF767422* JQ988220* KF537292* JQ988623* GQ395636* GQ141571 GQ395658* GQ141571 KF537315* JN819379* JN788938* KM211351* MF077654 KF537283 KF537297* KF537298 KF537308 JQ988527* KJ661260

ro

Haplogroups

369

Colombia

376 377

Peru Ecuador

38

Journal Pre-proof

380

Peru

446

Peru

464

Peru

481 499 500 502

Colombia Colombia Costa Rica Colombia Ecuador

re

lP Ecuador

Jo

ur

560

Sur America Peru Peru Peru Peru Peru Peru Peru

na

509 510 513 515 518 520 550

-p

508

GenBank Accession JQ988404* JQ988206* JQ988414 KF767423 KU562245* JQ988255* JQ988257 KF537314* KF537327* JN819387* MF990727* JF833062 JF833063 JF833064 GU085190* JQ988516* JQ988508* JQ988544* KF767424* KF767418* AY172842* KU562249* JF833052* JF833053

of

Country Peru

ro

Haplogroups

39

Journal Pre-proof

Jo

ur

na

lP

re

-p

ro

of

Table B4. Genera and species of birds to calculated the S/G rate, this information was extracted from the data set of (Gil-Vargas and Sedano-Cruz, 2019). Cluster Country Locality Family Antilles NA Thraupidae Thraupidae Felixlândia Tyrannidae Brazil Palmas,Tocantins Thraupidae Vila Boa Thraupidae Cauca, PNN Gorgona, El Poblado Thraupidae Colombia Pereira Cardinalidae Bilsa Fringillidae Loma Alta Tyrannidae Fringillidae Ecuador Provincia de Morona Santiago, Wisui Thraupidae 4 Tiputini Fringillidae Wisui reserve Thraupidae Nuevo Mexico, El Malpais Cardinalidae United States Nuevo Mexico, Mesa Chivato Cardinalidae Cardinalidae Guiana NA Icteridae Thraupidae Continental North America NA Cardinalidae Icteridae Uruguay NA Picidae Turdidae Thraupidae Scolopacidae Cauca, PNN Gorgona, El Poblado Thamnophilidae 39 Colombia Thraupidae Tyrannidae Quindío, Quimbaya, Reserva Natural La montaña del Ocaso Thamnophilidae 502 Colombia Cauca, PNN Gorgona, El Poblado Thraupidae

40

Genus Coereb Nemos Hemitr Tangar Tangar Coereb Piranga Euphon Mione Euphon Saltato Euphon Saltato Piranga Piranga Caryoth Psaroc Saltato

Piranga Pseudo Colapte Turdus Cyaner Actitis Thamn Coereb Tyrann

Thamn Sporop

Journal Pre-proof Table B5. Descriptive statistics and nucleotide variation for 78 haplogroups of Haemoproteus. The comparison between Gorgona Island and nearby areas of the Eastern Tropical Pacific includes the following avian groupings: resident, migratory, marine and shorebirds. Variable sites of cyt b gene (s), the richness of haplotype groups (h), expected heterozygosity (He), nucleotide diversity (π) and the Tajima´s D.

Resident birds

π (+/- s.d.)

Tajima´s D

P-value

33

8

0.02916

0.028125 +/- 0.015335

0.9823

0.8744

78

9

0.07344

0.071338 +/- 0.035455

2.1188

0.9877

98

26

0.03536

0.035455 +/- 0.017823

-0.859

0.2024

15

3

0.00611

0.006111 +/- 0.003722

-1.0808

0.1419

ur

na

lP

re

-p

birds of

He

of

Migratory birds Marine and

h

Jo

Haemoproteus in Haemoproteus in Shorebirds Haemoproteus in to mainland Haemoproteus in Gorgona Island

s

ro

Avian Groups

41

Journal Pre-proof Highlights Prevalence on Gorgona Island is higher than any local report in the Americas. Specific host-species effect of avian Haemosporidia to Gorgona Island. An avian Haemoproteus might be endemic to Gorgona Island.

Jo

ur

na

lP

re

-p

ro

of

Haemoproteus on Gorgona Island is similar to those circulating in the Americas.

42