Apparent Absence of Avian Malaria and Malaria-Like Parasites in Northern Blue-Footed Boobies Breeding On Isla Isabel

Apparent Absence of Avian Malaria and Malaria-Like Parasites in Northern Blue-Footed Boobies Breeding On Isla Isabel | Research Square window.SnipcartSettings = { analytics: { enabled: false } }; (function() { var accessVector = localStorage.getItem('access_vector') || ''; window.dataLayer = window.dataLayer || []; if (accessVector) { window.dataLayer.push({ user: { profile: { profileInfo: { snid: accessVector } } } }); } })(); (function(w,d,s,l,i){w[l]=w[l]||[];w[l].push({'gtm.start':new Date().getTime(),event:'gtm.js'});var f=d.getElementsByTagName(s)[0],j=d.createElement(s),dl=l!='dataLayer'?'&l='+l:'';j.async=true;j.src='https://www.googletagmanager.com/gtm.js?id='+i+dl;f.parentNode.insertBefore(j,f);})(window,document,'script','dataLayer','GTM-K279D39R'); Browse Preprints In Review Journals COVID-19 Preprints AJE Video Bytes Research Tools Research Promotion AJE Professional Editing AJE Rubriq About Preprint Platform In Review Editorial Policies Our Team Advisory Board Help Center Sign In Submit a Preprint Cite Share Download PDF Research Article Apparent Absence of Avian Malaria and Malaria-Like Parasites in Northern Blue-Footed Boobies Breeding On Isla Isabel Federico Roldán-Zurabián, María José Ruiz-López, Josué Martínez de la Puente, and 3 more This is a preprint; it has not been peer reviewed by a journal. https://doi.org/ 10.21203/rs.3.rs-842608/v1 This work is licensed under a CC BY 4.0 License Status: Posted Version 1 posted You are reading this latest preprint version Abstract Haemosporidian parasites are common in birds, but often are not in seabirds. The absence of vectors/genetic resistance to infection have been proposed to explain this pattern. Examination of different host populations is required to confirm the absence of blood parasites in widespread host species, which could be differently exposed to blood parasites across their geographic range. Moreover, screening of blood parasites in many seabirds has been done only by visual inspection of blood smears, which can miss low-intensity infections. Screening of blood parasites of the genera Plasmodium , Haemoproteus and Leucocytozoon , combining inspection of blood smears and PCR-based detection methods, revealed that a highly philopatric colony of blue-footed boobies ( Sula nebouxii ) in the Tropical North Pacific is likely free of these parasites. Earlier detection of Haemoproteus parasites in frigatebirds cohabiting with boobies in our study site and blue-footed boobies breeding on the Galapagos Islands suggests that absence of blood parasites in this northern booby colony could not be attributable to the absence of vectors or genetic resistance to infection. High host specificity or fine-scale spatial heterogeneity in the abundance of insect vectors could explain our negative results, but these hypotheses remain to be tested. We emphasize the relevance of assessing the occurrence of blood parasites in different populations of widespread host species, such as blue-footed boobies. Agroecology Epidemiology Avian malaria Plasmodium Haemoproteus Leucocytozoon Wild birds Seabirds Introduction Plasmodium , Haemoproteus , and Leucocytozoon blood parasites are widespread parasites causing avian malaria and avian malaria–like diseases 1 that have deleterious impacts on wild birds 2 – 4 . However, infections by these parasites are not homogeneously distributed among bird taxa, with some groups, including seabirds, usually showing an extremely low prevalence or total absence of parasite infections 5 . Different hypotheses have been proposed to explain these patterns including that absence of parasites is the result of adverse conditions for vectors in seabird habitats 6 . Seabird habitats usually have high salinity, high wind exposure and low vegetation cover, which are expected to impose strong selective pressures on vectors making them infrequent 6 , 7 . But even in the co-occurrence of potential vectors, blood parasite infections in seabirds are still uncommon 5 , suggesting that alternative hypotheses could also explain the observed patterns. Scarcity of infection in seabirds could also be due to good immunological capabilities of the hosts to repel infections, short exposure times to parasitic infections, or lack of suitable host-parasite assemblages 5 , 8 , 9 . Blue-footed boobies are socially monogamous seabirds 10 that breed colonially on islands of the Eastern Tropical Pacific Ocean, from Mexico’s Gulf of California to northern Peru 11 . Birds nest on the ground in open sites or in areas with moderate vegetation cover 11 . Isla Isabel boobies show lifetime fidelity to their first breeding site in their natal colony 10 , 12 and may live 20 years or longer 13 . Previous studies on this species revealed an approximately 33–83% prevalence of Haemoproteus sp. on the Galapagos islands 14 , implying physiological compatibility between birds and blood parasites. These infections were only detected when using molecular tools; in blood smears only a parasite tentatively identified as Leucocytozoon sp. was found. Here, we combined microscopic inspection of blood smears and molecular screening to assess the prevalence, and eventually the genetic identification, of blood parasites belonging to the genera Plasmodium , Haemoproteus and Leucocytozoon in adult blue-footed boobies breeding in an insular and highly philopatric colony on Isla Isabel (Mexico). We expect Isla Isabel boobies to be infected by blood parasites for two reasons. First, several species of mosquitoes belonging to the genera Aedes , Anopheles , Culex and the potential vectors of Haemoproteus and Leucocytozoon , including Culicoides , hippoboscids and black flies 15 , are widely distributed in Mexico and have been recorded in the study area ( 16 , 17 , authors per. obs.). Second, the prevalence of Haemoproteus iwa in Isla Isabel frigatebirds, likely vectored by hippoboscid flies 18 , has been estimated to be 16% (n = 251 frigatebirds; 19 ), reinforcing the idea that there are suitable insect vectors of haemoparasites on the island. Results A total of 64 blood smears from 33 male and 31 female blue-footed boobies were visually inspected for the presence of blood parasites. No parasites were detected in blood smears. None of the 64 samples of blue-booted boobies tested molecularly showed evidence of parasite amplification. Discussion The inspection of Haemosporidian parasites of the genera Plasmodium , Haemoproteus and Leucocytozoon , combining microscopic screening of blood smears and a broadly used PCR-based detection method, suggests that adult male and female blue-footed boobies breeding on Isla Isabel are likely free of these blood parasites. Our results support previous findings by Clark and Swinehart based on blood smears 20 who recorded the absence of these parasites in 19 blue-footed boobies from islands off the North Pacific coast of Mexico (not including Isla Isabel boobies). These negative results reflect the overall scarcity of blood parasites in seabirds, which is often attributed to unsuitable conditions for vectors in seabird habitats 5 , 21 . In addition to the occurrence of potential vectors in the area, Haemoproteus parasites have been previously detected in frigatebirds from Isla Isabel (prevalence ranging from 16 to 54%; 19 , 22 ), thus lack of blood parasites in Isla Isabel boobies probably implies that parasite infections vary among avian host species that coexist on the island. Differences in the prevalence of blood parasites between boobies and frigatebirds could be due to differences in their habitat use or in the abundance and activity of vectors 23 , 24 . Infected frigatebirds were sampled in the southwestern end of the island, in a shady, vegetated area with large trees located a few ten of meters from an unfinished building where rainwater accumulates, and 150 meters from the only pool of alkaline water on the island 19 , 25 . This area is protected from the wind and suitable for development and proliferation of some potential vectors 26 . In addition, the parasites detected in frigatebirds are transmitted by louse flies (Hippoboscidae) that spend most of the time on their vertebrate hosts and present higher host specificity than mosquitoes, Culicoides and black flies. Boobies breed mostly where we sampled them in the wind-exposed northeastern end of the island, ~ 1.0–1.5 km from the source of alkaline water pool and the unfinished building where rainwater accumulates. The northeastern area is mainly covered by stunted woody vegetation, since big trees have been recurrently damaged by hurricanes 27 , 28 . Windy, dry, and hot conditions limit the abundance and activity of potential vectors of blood parasites in marine environments 6 , 23 , and their prevalence in the Isla Isabel booby colony may explain the absence of blood parasites in boobies. Lack of blood parasites in seabirds has been attributed to evolutionary and ecological factors other than spatial heterogeneity in the occurrence of suitable vectors 5 , 8 , but it is unlikely that these alternative mechanisms explain the results reported here. Earlier screening of blood parasites in boobies from the Galápagos archipelago revealed the infection by a tentatively identified Leucocytozoon parasite, and especially, by parasites of Haemoproteus genus 14 , 18 . Thus, it is unlikely that blood parasites are absent from Isla Isabel boobies because they exhibit high immunological capacities, have short exposure times to parasitic infections or present physiological incompatibility with haemoparasites that prevents the latter from completing their life cycles 8 , 9 . In addition, differences in host numbers 8 may not explain differences in the occurrence of blood parasites between the booby populations of Isla Isabel and the Galápagos. The booby colony of Isla Isabel has up to 1769 breeding pairs in our study area alone, which covers 26, 889 m 2 and contains approximately 65% of all breeding pairs that are established annually on the island 29 . The breeding population of blue-footed boobies in the whole Galápagos archipelago was estimated at 3,200 pairs at the time when high prevalence of blood parasites was detected in Galápagos boobies 14 , 30 . Thus, host numbers are not different enough between the two populations to explain the apparent lack of blood parasites in Isla Isabel boobies. In conclusion, we failed to find evidence of blood parasites in one of the largest colonies of blue-footed boobies of the North Pacific coast, after microscopic examination of blood smears and state-of-the-art molecular analysis for detection of avian blood parasites 9 , 31 . Apparent absence of blood parasites in Isla Isabel boobies indirectly adds to the growing evidence of variation in parasite infections among avian host species that coexist locally 23 , 32 , and highlights the relevance of performing evaluations of the prevalence of blood parasites in different populations of widespread host species 21 . Methods Study site Isla Isabel is an 82-ha volcanic island 28 km off the west coast of Mexico, in the Eastern Tropical North Pacific (21º 52´ N, 105º 54´W). The island is mainly covered by deciduous dry forest of Crataeva tapia trees, Euphorbia schlechtendalli bushes, and an understory dominated by coastal grasslands. The climate is sub-humid tropical with rains in June–November (the hurricane season). In the rainy season, water levels rise both on the steep rocky slopes and valleys, and water is collected in three endorheic basins in the center and north of the island. There is also a shallow pool of alkaline water of approximately 50 m in diameter in the south of the island that is the result of rainwater runoff and the entry of seawater during storms 27 . Field procedures In March-April 2019, we hand-captured 64 adult blue-footed boobies (33 males and 31 females) on their nests (sites with a clutch or brood) at night. We recorded the identity of boobies that had a metal ring (since 1989, tens of thousands of fledglings and adults have been banded on Isla Isabel 13 ) and sexed by voice (females grunt, males whistle). See Drummond et al. 33 for further details on the field procedures used. We blood sampled all captured boobies only once. Approximately 1.5–2.0 ml of blood were taken from their brachial vein. A drop of blood was used for blood smear preparation and 200 µl were split in two aliquots (100 µl each) and stored in 96% ethanol for molecular screening of blood parasites. The remaining blood was centrifuged and stored for future research. Blood smears were fixed in 96% ethanol and subsequently stained with Giemsa. Manipulation of booby adults took less than 10 min and bleeding stopped before release at the site of capture. All adults resumed nest attendance 5–10 min after release. Data collection and blood sampling protocols comply with the current laws and ethical standards of animal research in Mexico (NOM-059-SEMARNAT-2010) and were revised and approved by the Secretaría de Medio Ambiente y Recursos Naturales (SEMARNAT; permit number SGPA/DGVS/01216617). In addition, we confirm that all methods are reported in accordance with ARRIVE guidelines 2.0 ( https://arriveguidelines.org ). Parasite identification Blood smears were scanned for the presence of blood parasites using a light microscope Nikon Eclipse Ti - Arcturus XT of Applied Biosystems. Half of each blood smear was scanned at 400x magnification in search of larger parasites, including Leucocytozoon , during 30–45 min. The other half of each smear was scanned at 1000x magnification in search of Haemoproteus , Plasmodium and Leucocytozoon parasites in up to > 10,000 erythrocytes per smear (see 34 ) during 30 min. DNA from blood samples were extracted using the Maxwell®16 LEV system Research (Promega, Madison, WI) 35 . Samples were run in 0.8% agarose gels to check DNA integrity. To detect and identify avian parasites of the genera Plasmodium , Haemoproteus and Leucocytozoon , we carried out nested PCRs following Hellgren et al. 36 to amplify a fragment of the mitochondrial cytochrome b gene. To check if the PCRs amplified successfully, we ran 3.0 µl of the final PCR product on a 2% agarose gel. We ran 6 repetitions of each PCR to exclude false negatives. Declarations Acknowledgements We thank Karla Guerrero, Daniel Linke and Carrie Gunn for their assistance in the field, the Secretaría de Marina and fishermen from Nayarit for logistical support, and the SEMARNAT and CONANP for permission to work on Isla Isabel. Cristina Rodríguez provided logistical support, Gastón Contreras-Jiménez provided technical assistance in microscopy analysis, and Isabel Martín and Cristina Perez provided technical assistance in the laboratory. Author contributions SA and HD conceived the study. FRZ and SA sampled birds. FRZ, JMP, MJRL and JF analyzed the samples. SA drafted the manuscript with significant contributions of the other authors. All authors read and approved the manuscript. Competing interests The authors declare no competing interests. Data availability Data supporting the conclusions are included within the article. Ethics approval Our research complies with Mexican legal and ethical requirements. Permit to collect blood samples (SGPA/DGVS/01216617) was provided by Secretaría del Medio Ambiente y Recursos Naturales (SEMARNAT). Consent for publication Not applicable. References Atkinson, C. T. & Van Riper, C. Pathogenicity and epizootiology of avian haematozoa: Plasmodium, Leucocytozoon, and Haemoproteus. Bird-parasite Interact. 19–48(1991). Sorci, G. & Moller, A. P. Comparative evidence for a positive correlation between haematozoan prevalence and mortality in waterfowl. J. Evol. Biol , 10 , 731–741 (1997). Merino, S., Moreno, J., Sanz, J. J. & Arriero, E. Are avian blood parasites pathogenic in the wild? A medication experiment in blue tits (Parus caeruleus). Proc. R. Soc. B Biol. Sci. 267, 2507–2510(2000). Asghar, M. et al. Hidden costs of infection: Chronic malaria accelerates telomere degradation and senescence in wild birds. Science (80-.) , 347 , 436–438 (2015). Quillfeldt, P., Arriero, E., Martínez, J., Masello, J. F. & Merino, S. Prevalence of blood parasites in seabirds - a review. Front. Zool , 8 , 26 (2011). Piersma, T. Do Global Patterns of Habitat Use and Migration Strategies Co-Evolve with Relative Investments in Immunocompetence due to Spatial Variation in Parasite Pressure?, 80 , 623 (1997). Mendes, L., Piersma, T., Lecoq, M., Spaans, B. & Ricklefs, R. E. Disease-limited distributions? Contrasts in the prevalence of avian malaria in shorebird species using marine and freshwater habitats., 109 , 396–404 (2005). Martínez-Abraín, A., Esparza, B. & Oro, D. Lack of blood parasites in bird species: Does absence of blood parasite vectors explain it all? in Ardeola vol. 51 225–232(2004). Campioni, L. et al. Absence of haemosporidian parasite infections in the long-lived Cory’s shearwater: evidence from molecular analyses and review of the literature. Parasitol. Res , 117 , 323–329 (2018). Osorio-Beristain, M. & Drummond, H. Non-aggressive mate guarding by the blue-footed booby: A balance of female and male control. Behav. Ecol. Sociobiol , 43 , 307–315 (1998). Nelson, J. B. Pelicans, cormorants and their relatives: The Pelecaniformes (Oxford University Press, 2006). Kim, S. Y., Torres, R., Domínguez, C. A. & Drummond, H. Lifetime philopatry in the blue-footed booby: A longitudinal study. Behav. Ecol , 18 , 1132–1138 (2007). Drummond, H. & Rodríguez, C. Viability of booby offspring is maximized by having one young parent and one old parent. PLoS One , 10 , e0133213 (2015). Lee-Cruz, L. et al. Prevalence of Haemoproteus sp. in Galápagos blue-footed boobies: effects on health and reproduction. Parasitol. Open 2 , (2016). Santiago-Alarcon, D., Palinauskas, V. & Schaefer, H. M. Diptera vectors of avian Haemosporidian parasites: Untangling parasite life cycles and their taxonomy. Biol. Rev , 87 , 928–964 (2012). Bond, J. G. et al. Diversity of mosquitoes and the aquatic insects associated with their oviposition sites along the Pacific coast of Mexico. Parasites and Vectors , 7 , 1–19 (2014). Ibañez-Bernal, S. Informe Final del proyecto Actualización del Catálogo de Autoridad Taxonómica del orden Diptera (Insecta) de México CONABIO (JE006)(2017). Levin, I. I. et al. Hippoboscid-transmitted Haemoproteus parasites (Haemosporida) infect Galapagos Pelecaniform birds: Evidence from molecular and morphological studies, with a description of Haemoproteus iwa. Int. J. Parasitol , 41 , 1019–1027 (2011). Madsen, V. et al. Testosterone levels and gular pouch coloration in courting magnificent frigatebird (Fregata magnificens): Variation with age-class, visited status and blood parasite infection. Horm. Behav , 51 , 156–163 (2007). Clark, G. W. & Swinehart, B. Avian haematozoa from the offshore islands of northern Mexico. Wildl. Dis , 5 , 111–112 (1969). Quillfeldt, P. et al. Hemosporidian blood parasites in seabirds - A comparative genetic study of species from Antarctic to tropical habitats., 97 , 809–817 (2010). Merino, S. et al. Infection by haemoproteus parasites in four species of frigatebirds and the description of a new species of Haemoproteus (Haemosporida: Haemoproteidae). J. Parasitol , 98 , 388–397 (2012). Svensson, L. M. E. & Ricklefs, R. E. Low diversity and high intra-island variation in prevalence of avian Haemoproteus parasites on Barbados, Lesser Antilles., 136 , 1121–1131 (2009). Loiseau, C. et al. Spatial variation of haemosporidian parasite infection in african rainforest bird species. J. Parasitol , 96 , 21–29 (2010). Madsen, V. Female mate choice in the magnificent frigatebird (Fregata magnificens) (Universidad Nacional Autónoma de México, 2004). Super, P. E. & van Riper, C. A comparison of avian hematozoan epizootiology in two California coastal scrub communities. J. Wildl. Dis , 31 , 447–461 (1995). CONANP. Programa de Conservación y Manejo del Parque Nacional Isla Isabel . (2005). Ancona, S., Drummond, H., Rodríguez, C. & Zúñiga-Vega, J. J. Long-term population dynamics reveal that survival and recruitment of tropical boobies improve after a hurricane. J. Avian Biol , 48 , 320–332 (2017). Ancona, S., Sánchez-Colón, S., Rodríguez, C. & Drummond, H. El Niño in the Warm Tropics: Local sea temperature predicts breeding parameters and growth of blue-footed boobies. J. Anim. Ecol , 80 , 799–808 (2011). Anderson, D. J., Huyvaert, K. P. & Anchundia, D. Chronic lack of breeding by Galápagos Blue-footed Boobies and associated population decline. Galapagos Report 2013 –2014 vol. 9 https://www.galapagos.org/wp-content/uploads/2015/09/GalapagosReport_2013-2014-22-Anderson-158-163.pdf (2015). Valkiunas, G. et al. A comparative analysis of microscopy and PCR-based detection methods for blood parasites. J. Parasitol , 94 , 1395–1401 (2008). Santiago-Alarcon, D. et al. Parasites in space and time: a case study of haemosporidian spatiotemporal prevalence in urban birds. Int. J. Parasitol , 49 , 235–246 (2019). Drummond, H., Torres, R. & Krishnan, V. V. Buffered development: Resilience after aggressive subordination in infancy. Am. Nat , 161 , 794–807 (2003). Merino, S. & Potti, J. High Prevalence of Hematozoa in Nestlings of a Passerine Species, the Pied Flycatcher (Ficedula hypoleuca). Auk , 112 , 1041–1043 (1995). Gutiérrez-López, R. et al. Low prevalence of blood parasites in a long-distance migratory raptor: The importance of host habitat. Parasites and Vectors , 8 , 1–6 (2015). Hellgren, O., Waldenström, J. & Bensch, S. A new PCR assay for simultaneous studies of Leucocytozoon, Plasmodium, and Haemoproteus from avian blood. J. Parasitol , 90 , 797–802 (2004). Additional Declarations No competing interests reported. Cite Share Download PDF Status: Posted Version 1 posted You are reading this latest preprint version Research Square lets you share your work early, gain feedback from the community, and start making changes to your manuscript prior to peer review in a journal. As a division of Research Square Company, we’re committed to making research communication faster, fairer, and more useful. We do this by developing innovative software and high quality services for the global research community. Our growing team is made up of researchers and industry professionals working together to solve the most critical problems facing scientific publishing. Also discoverable on Platform About Our Team In Review Editorial Policies Advisory Board Help Center Resources Author Services Accessibility API Access RSS feed Manage Cookie Preferences © Research Square 2026 | ISSN 2693-5015 (online) Privacy Policy Terms of Service Do Not Sell My Personal Information {"props":{"pageProps":{"initialData":{"identity":"rs-842608","acceptedTermsAndConditions":true,"allowDirectSubmit":true,"archivedVersions":[],"articleType":"Research Article","associatedPublications":[],"authors":[{"id":48157508,"identity":"14431dd5-0f0f-40e4-a48d-fccaeade0bb5","order_by":0,"name":"Federico Roldán-Zurabián","email":"","orcid":"","institution":"National Autonomous University of Mexico","correspondingAuthor":false,"prefix":"","firstName":"Federico","middleName":"","lastName":"Roldán-Zurabián","suffix":""},{"id":48157510,"identity":"7436a2f5-a592-46d6-96d3-75051d3ab9fb","order_by":1,"name":"María José Ruiz-López","email":"","orcid":"","institution":"Estación Biológica de Doñana","correspondingAuthor":false,"prefix":"","firstName":"María","middleName":"José","lastName":"Ruiz-López","suffix":""},{"id":48157512,"identity":"954b8a7a-1556-4fc9-a4ca-3f4ad658f2a4","order_by":2,"name":"Josué Martínez de la Puente","email":"","orcid":"","institution":"University of Granada","correspondingAuthor":false,"prefix":"","firstName":"Josué","middleName":"Martínez de la","lastName":"Puente","suffix":""},{"id":48157513,"identity":"5718983b-ee57-4e47-b8a4-1337bec9e3c9","order_by":3,"name":"Jordi Figuerola","email":"","orcid":"","institution":"Estación Biológica de Doñana","correspondingAuthor":false,"prefix":"","firstName":"Jordi","middleName":"","lastName":"Figuerola","suffix":""},{"id":48157514,"identity":"95f0a2f4-56e5-4349-8f81-85fa8f122eb8","order_by":4,"name":"Hugh Drummond","email":"","orcid":"","institution":"National Autonomous University of Mexico","correspondingAuthor":false,"prefix":"","firstName":"Hugh","middleName":"","lastName":"Drummond","suffix":""},{"id":48157515,"identity":"6cff8f94-153b-4995-bc79-64916ff54076","order_by":5,"name":"Sergio Ancona","email":"data:image/png;base64,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","orcid":"","institution":"National Autonomous University of Mexico","correspondingAuthor":true,"prefix":"","firstName":"Sergio","middleName":"","lastName":"Ancona","suffix":""}],"badges":[],"createdAt":"2021-08-24 22:44:03","currentVersionCode":1,"declarations":"","doi":"10.21203/rs.3.rs-842608/v1","doiUrl":"https://doi.org/10.21203/rs.3.rs-842608/v1","draftVersion":[],"editorialEvents":[],"editorialNote":"","failedWorkflow":false,"files":[{"id":15907399,"identity":"71c3ddc6-87f0-4a0f-9b33-f834337146a1","added_by":"auto","created_at":"2021-11-26 07:14:16","extension":"pdf","order_by":0,"title":"","display":"","copyAsset":false,"role":"manuscript-pdf","size":238727,"visible":true,"origin":"","legend":"","description":"","filename":"manuscript.pdf","url":"https://assets-eu.researchsquare.com/files/rs-842608/v1/76f105da-c9af-4929-af73-1e6f791e0664.pdf"}],"financialInterests":"No competing interests reported.","formattedTitle":"\u003cp\u003eApparent Absence of Avian Malaria and Malaria-Like Parasites in Northern Blue-Footed Boobies Breeding On Isla Isabel\u003c/p\u003e","fulltext":[{"header":"Introduction","content":"\u003cp\u003e \u003cem\u003ePlasmodium\u003c/em\u003e, \u003cem\u003eHaemoproteus\u003c/em\u003e, and \u003cem\u003eLeucocytozoon\u003c/em\u003e blood parasites are widespread parasites causing avian malaria and avian malaria\u0026ndash;like diseases \u003csup\u003e\u003cspan citationid=\"CR1\" class=\"CitationRef\"\u003e1\u003c/span\u003e\u003c/sup\u003e that have deleterious impacts on wild birds \u003csup\u003e\u003cspan additionalcitationids=\"CR3\" citationid=\"CR2\" class=\"CitationRef\"\u003e2\u003c/span\u003e\u0026ndash;\u003cspan citationid=\"CR4\" class=\"CitationRef\"\u003e4\u003c/span\u003e\u003c/sup\u003e. However, infections by these parasites are not homogeneously distributed among bird taxa, with some groups, including seabirds, usually showing an extremely low prevalence or total absence of parasite infections \u003csup\u003e\u003cspan citationid=\"CR5\" class=\"CitationRef\"\u003e5\u003c/span\u003e\u003c/sup\u003e. Different hypotheses have been proposed to explain these patterns including that absence of parasites is the result of adverse conditions for vectors in seabird habitats \u003csup\u003e\u003cspan citationid=\"CR6\" class=\"CitationRef\"\u003e6\u003c/span\u003e\u003c/sup\u003e. Seabird habitats usually have high salinity, high wind exposure and low vegetation cover, which are expected to impose strong selective pressures on vectors making them infrequent \u003csup\u003e\u003cspan citationid=\"CR6\" class=\"CitationRef\"\u003e6\u003c/span\u003e,\u003cspan citationid=\"CR7\" class=\"CitationRef\"\u003e7\u003c/span\u003e\u003c/sup\u003e. But even in the co-occurrence of potential vectors, blood parasite infections in seabirds are still uncommon \u003csup\u003e\u003cspan citationid=\"CR5\" class=\"CitationRef\"\u003e5\u003c/span\u003e\u003c/sup\u003e, suggesting that alternative hypotheses could also explain the observed patterns. Scarcity of infection in seabirds could also be due to good immunological capabilities of the hosts to repel infections, short exposure times to parasitic infections, or lack of suitable host-parasite assemblages \u003csup\u003e\u003cspan citationid=\"CR5\" class=\"CitationRef\"\u003e5\u003c/span\u003e,\u003cspan citationid=\"CR8\" class=\"CitationRef\"\u003e8\u003c/span\u003e,\u003cspan citationid=\"CR9\" class=\"CitationRef\"\u003e9\u003c/span\u003e\u003c/sup\u003e.\u003c/p\u003e \u003cp\u003eBlue-footed boobies are socially monogamous seabirds \u003csup\u003e\u003cspan citationid=\"CR10\" class=\"CitationRef\"\u003e10\u003c/span\u003e\u003c/sup\u003e that breed colonially on islands of the Eastern Tropical Pacific Ocean, from Mexico\u0026rsquo;s Gulf of California to northern Peru \u003csup\u003e\u003cspan citationid=\"CR11\" class=\"CitationRef\"\u003e11\u003c/span\u003e\u003c/sup\u003e. Birds nest on the ground in open sites or in areas with moderate vegetation cover \u003csup\u003e\u003cspan citationid=\"CR11\" class=\"CitationRef\"\u003e11\u003c/span\u003e\u003c/sup\u003e. Isla Isabel boobies show lifetime fidelity to their first breeding site in their natal colony \u003csup\u003e\u003cspan citationid=\"CR10\" class=\"CitationRef\"\u003e10\u003c/span\u003e,\u003cspan citationid=\"CR12\" class=\"CitationRef\"\u003e12\u003c/span\u003e\u003c/sup\u003e and may live 20 years or longer \u003csup\u003e\u003cspan citationid=\"CR13\" class=\"CitationRef\"\u003e13\u003c/span\u003e\u003c/sup\u003e. Previous studies on this species revealed an approximately 33\u0026ndash;83% prevalence of \u003cem\u003eHaemoproteus\u003c/em\u003e sp. on the Galapagos islands \u003csup\u003e\u003cspan citationid=\"CR14\" class=\"CitationRef\"\u003e14\u003c/span\u003e\u003c/sup\u003e, implying physiological compatibility between birds and blood parasites. These infections were only detected when using molecular tools; in blood smears only a parasite tentatively identified as \u003cem\u003eLeucocytozoon\u003c/em\u003e sp. was found.\u003c/p\u003e \u003cp\u003eHere, we combined microscopic inspection of blood smears and molecular screening to assess the prevalence, and eventually the genetic identification, of blood parasites belonging to the genera \u003cem\u003ePlasmodium\u003c/em\u003e, \u003cem\u003eHaemoproteus\u003c/em\u003e and \u003cem\u003eLeucocytozoon\u003c/em\u003e in adult blue-footed boobies breeding in an insular and highly philopatric colony on Isla Isabel (Mexico). We expect Isla Isabel boobies to be infected by blood parasites for two reasons. First, several species of mosquitoes belonging to the genera \u003cem\u003eAedes\u003c/em\u003e, \u003cem\u003eAnopheles\u003c/em\u003e, \u003cem\u003eCulex\u003c/em\u003e and the potential vectors of \u003cem\u003eHaemoproteus\u003c/em\u003e and \u003cem\u003eLeucocytozoon\u003c/em\u003e, including \u003cem\u003eCulicoides\u003c/em\u003e, hippoboscids and black flies \u003csup\u003e\u003cspan citationid=\"CR15\" class=\"CitationRef\"\u003e15\u003c/span\u003e\u003c/sup\u003e, are widely distributed in Mexico and have been recorded in the study area (\u003csup\u003e\u003cspan citationid=\"CR16\" class=\"CitationRef\"\u003e16\u003c/span\u003e,\u003cspan citationid=\"CR17\" class=\"CitationRef\"\u003e17\u003c/span\u003e\u003c/sup\u003e, authors per. obs.). Second, the prevalence of \u003cem\u003eHaemoproteus iwa\u003c/em\u003e in Isla Isabel frigatebirds, likely vectored by hippoboscid flies \u003csup\u003e\u003cspan citationid=\"CR18\" class=\"CitationRef\"\u003e18\u003c/span\u003e\u003c/sup\u003e, has been estimated to be 16% (n\u0026thinsp;=\u0026thinsp;251 frigatebirds; \u003csup\u003e\u003cspan citationid=\"CR19\" class=\"CitationRef\"\u003e19\u003c/span\u003e\u003c/sup\u003e), reinforcing the idea that there are suitable insect vectors of haemoparasites on the island.\u003c/p\u003e"},{"header":"Results","content":"\u003cp\u003eA total of 64 blood smears from 33 male and 31 female blue-footed boobies were visually inspected for the presence of blood parasites. No parasites were detected in blood smears. None of the 64 samples of blue-booted boobies tested molecularly showed evidence of parasite amplification.\u003c/p\u003e"},{"header":"Discussion","content":"\u003cp\u003eThe inspection of Haemosporidian parasites of the genera \u003cem\u003ePlasmodium\u003c/em\u003e, \u003cem\u003eHaemoproteus\u003c/em\u003e and \u003cem\u003eLeucocytozoon\u003c/em\u003e, combining microscopic screening of blood smears and a broadly used PCR-based detection method, suggests that adult male and female blue-footed boobies breeding on Isla Isabel are likely free of these blood parasites. Our results support previous findings by Clark and Swinehart based on blood smears \u003csup\u003e\u003cspan citationid=\"CR20\" class=\"CitationRef\"\u003e20\u003c/span\u003e\u003c/sup\u003e who recorded the absence of these parasites in 19 blue-footed boobies from islands off the North Pacific coast of Mexico (not including Isla Isabel boobies). These negative results reflect the overall scarcity of blood parasites in seabirds, which is often attributed to unsuitable conditions for vectors in seabird habitats \u003csup\u003e\u003cspan citationid=\"CR5\" class=\"CitationRef\"\u003e5\u003c/span\u003e,\u003cspan citationid=\"CR21\" class=\"CitationRef\"\u003e21\u003c/span\u003e\u003c/sup\u003e.\u003c/p\u003e \u003cp\u003eIn addition to the occurrence of potential vectors in the area, \u003cem\u003eHaemoproteus\u003c/em\u003e parasites have been previously detected in frigatebirds from Isla Isabel (prevalence ranging from 16 to 54%; \u003csup\u003e\u003cspan citationid=\"CR19\" class=\"CitationRef\"\u003e19\u003c/span\u003e,\u003cspan citationid=\"CR22\" class=\"CitationRef\"\u003e22\u003c/span\u003e\u003c/sup\u003e), thus lack of blood parasites in Isla Isabel boobies probably implies that parasite infections vary among avian host species that coexist on the island. Differences in the prevalence of blood parasites between boobies and frigatebirds could be due to differences in their habitat use or in the abundance and activity of vectors \u003csup\u003e\u003cspan citationid=\"CR23\" class=\"CitationRef\"\u003e23\u003c/span\u003e,\u003cspan citationid=\"CR24\" class=\"CitationRef\"\u003e24\u003c/span\u003e\u003c/sup\u003e. Infected frigatebirds were sampled in the southwestern end of the island, in a shady, vegetated area with large trees located a few ten of meters from an unfinished building where rainwater accumulates, and 150 meters from the only pool of alkaline water on the island \u003csup\u003e\u003cspan citationid=\"CR19\" class=\"CitationRef\"\u003e19\u003c/span\u003e,\u003cspan citationid=\"CR25\" class=\"CitationRef\"\u003e25\u003c/span\u003e\u003c/sup\u003e. This area is protected from the wind and suitable for development and proliferation of some potential vectors \u003csup\u003e\u003cspan citationid=\"CR26\" class=\"CitationRef\"\u003e26\u003c/span\u003e\u003c/sup\u003e. In addition, the parasites detected in frigatebirds are transmitted by louse flies (Hippoboscidae) that spend most of the time on their vertebrate hosts and present higher host specificity than mosquitoes, \u003cem\u003eCulicoides\u003c/em\u003e and black flies. Boobies breed mostly where we sampled them in the wind-exposed northeastern end of the island, ~\u0026thinsp;1.0\u0026ndash;1.5 km from the source of alkaline water pool and the unfinished building where rainwater accumulates. The northeastern area is mainly covered by stunted woody vegetation, since big trees have been recurrently damaged by hurricanes \u003csup\u003e\u003cspan citationid=\"CR27\" class=\"CitationRef\"\u003e27\u003c/span\u003e,\u003cspan citationid=\"CR28\" class=\"CitationRef\"\u003e28\u003c/span\u003e\u003c/sup\u003e. Windy, dry, and hot conditions limit the abundance and activity of potential vectors of blood parasites in marine environments \u003csup\u003e\u003cspan citationid=\"CR6\" class=\"CitationRef\"\u003e6\u003c/span\u003e,\u003cspan citationid=\"CR23\" class=\"CitationRef\"\u003e23\u003c/span\u003e\u003c/sup\u003e, and their prevalence in the Isla Isabel booby colony may explain the absence of blood parasites in boobies.\u003c/p\u003e \u003cp\u003eLack of blood parasites in seabirds has been attributed to evolutionary and ecological factors other than spatial heterogeneity in the occurrence of suitable vectors \u003csup\u003e\u003cspan citationid=\"CR5\" class=\"CitationRef\"\u003e5\u003c/span\u003e,\u003cspan citationid=\"CR8\" class=\"CitationRef\"\u003e8\u003c/span\u003e\u003c/sup\u003e, but it is unlikely that these alternative mechanisms explain the results reported here. Earlier screening of blood parasites in boobies from the Gal\u0026aacute;pagos archipelago revealed the infection by a tentatively identified \u003cem\u003eLeucocytozoon\u003c/em\u003e parasite, and especially, by parasites of \u003cem\u003eHaemoproteus\u003c/em\u003e genus \u003csup\u003e\u003cspan citationid=\"CR14\" class=\"CitationRef\"\u003e14\u003c/span\u003e,\u003cspan citationid=\"CR18\" class=\"CitationRef\"\u003e18\u003c/span\u003e\u003c/sup\u003e. Thus, it is unlikely that blood parasites are absent from Isla Isabel boobies because they exhibit high immunological capacities, have short exposure times to parasitic infections or present physiological incompatibility with haemoparasites that prevents the latter from completing their life cycles \u003csup\u003e\u003cspan citationid=\"CR8\" class=\"CitationRef\"\u003e8\u003c/span\u003e,\u003cspan citationid=\"CR9\" class=\"CitationRef\"\u003e9\u003c/span\u003e\u003c/sup\u003e. In addition, differences in host numbers \u003csup\u003e\u003cspan citationid=\"CR8\" class=\"CitationRef\"\u003e8\u003c/span\u003e\u003c/sup\u003e may not explain differences in the occurrence of blood parasites between the booby populations of Isla Isabel and the Gal\u0026aacute;pagos. The booby colony of Isla Isabel has up to 1769 breeding pairs in our study area alone, which covers 26, 889 m\u003csup\u003e\u003cspan citationid=\"CR2\" class=\"CitationRef\"\u003e2\u003c/span\u003e\u003c/sup\u003e and contains approximately 65% of all breeding pairs that are established annually on the island \u003csup\u003e\u003cspan citationid=\"CR29\" class=\"CitationRef\"\u003e29\u003c/span\u003e\u003c/sup\u003e. The breeding population of blue-footed boobies in the whole Gal\u0026aacute;pagos archipelago was estimated at 3,200 pairs at the time when high prevalence of blood parasites was detected in Gal\u0026aacute;pagos boobies \u003csup\u003e\u003cspan citationid=\"CR14\" class=\"CitationRef\"\u003e14\u003c/span\u003e,\u003cspan citationid=\"CR30\" class=\"CitationRef\"\u003e30\u003c/span\u003e\u003c/sup\u003e. Thus, host numbers are not different enough between the two populations to explain the apparent lack of blood parasites in Isla Isabel boobies.\u003c/p\u003e \u003cp\u003eIn conclusion, we failed to find evidence of blood parasites in one of the largest colonies of blue-footed boobies of the North Pacific coast, after microscopic examination of blood smears and state-of-the-art molecular analysis for detection of avian blood parasites \u003csup\u003e\u003cspan citationid=\"CR9\" class=\"CitationRef\"\u003e9\u003c/span\u003e,\u003cspan citationid=\"CR31\" class=\"CitationRef\"\u003e31\u003c/span\u003e\u003c/sup\u003e. Apparent absence of blood parasites in Isla Isabel boobies indirectly adds to the growing evidence of variation in parasite infections among avian host species that coexist locally \u003csup\u003e\u003cspan citationid=\"CR23\" class=\"CitationRef\"\u003e23\u003c/span\u003e,\u003cspan citationid=\"CR32\" class=\"CitationRef\"\u003e32\u003c/span\u003e\u003c/sup\u003e, and highlights the relevance of performing evaluations of the prevalence of blood parasites in different populations of widespread host species \u003csup\u003e\u003cspan citationid=\"CR21\" class=\"CitationRef\"\u003e21\u003c/span\u003e\u003c/sup\u003e.\u003c/p\u003e"},{"header":"Methods","content":"\u003cdiv id=\"Sec5\" class=\"Section2\"\u003e \u003ch2\u003eStudy site\u003c/h2\u003e \u003cp\u003eIsla Isabel is an 82-ha volcanic island 28 km off the west coast of Mexico, in the Eastern Tropical North Pacific (21\u0026ordm; 52\u0026acute; N, 105\u0026ordm; 54\u0026acute;W). The island is mainly covered by deciduous dry forest of \u003cem\u003eCrataeva tapia\u003c/em\u003e trees, \u003cem\u003eEuphorbia schlechtendalli\u003c/em\u003e bushes, and an understory dominated by coastal grasslands. The climate is sub-humid tropical with rains in June\u0026ndash;November (the hurricane season). In the rainy season, water levels rise both on the steep rocky slopes and valleys, and water is collected in three endorheic basins in the center and north of the island. There is also a shallow pool of alkaline water of approximately 50 m in diameter in the south of the island that is the result of rainwater runoff and the entry of seawater during storms \u003csup\u003e\u003cspan citationid=\"CR27\" class=\"CitationRef\"\u003e27\u003c/span\u003e\u003c/sup\u003e.\u003c/p\u003e \u003c/div\u003e \u003cdiv id=\"Sec6\" class=\"Section2\"\u003e \u003ch2\u003eField procedures\u003c/h2\u003e \u003cp\u003eIn March-April 2019, we hand-captured 64 adult blue-footed boobies (33 males and 31 females) on their nests (sites with a clutch or brood) at night. We recorded the identity of boobies that had a metal ring (since 1989, tens of thousands of fledglings and adults have been banded on Isla Isabel \u003csup\u003e\u003cspan citationid=\"CR13\" class=\"CitationRef\"\u003e13\u003c/span\u003e\u003c/sup\u003e) and sexed by voice (females grunt, males whistle). See Drummond et al. \u003csup\u003e\u003cspan citationid=\"CR33\" class=\"CitationRef\"\u003e33\u003c/span\u003e\u003c/sup\u003e for further details on the field procedures used. We blood sampled all captured boobies only once. Approximately 1.5\u0026ndash;2.0 ml of blood were taken from their brachial vein. A drop of blood was used for blood smear preparation and 200 \u0026micro;l were split in two aliquots (100 \u0026micro;l each) and stored in 96% ethanol for molecular screening of blood parasites. The remaining blood was centrifuged and stored for future research. Blood smears were fixed in 96% ethanol and subsequently stained with Giemsa. Manipulation of booby adults took less than 10 min and bleeding stopped before release at the site of capture. All adults resumed nest attendance 5\u0026ndash;10 min after release. Data collection and blood sampling protocols comply with the current laws and ethical standards of animal research in Mexico (NOM-059-SEMARNAT-2010) and were revised and approved by the Secretar\u0026iacute;a de Medio Ambiente y Recursos Naturales (SEMARNAT; permit number SGPA/DGVS/01216617). In addition, we confirm that all methods are reported in accordance with ARRIVE guidelines 2.0 (\u003cspan class=\"ExternalRef\"\u003e\u003cspan class=\"RefSource\"\u003ehttps://arriveguidelines.org\u003c/span\u003e\u003c/span\u003e).\u003c/p\u003e \u003c/div\u003e \u003cdiv id=\"Sec7\" class=\"Section2\"\u003e \u003ch2\u003eParasite identification\u003c/h2\u003e \u003cp\u003eBlood smears were scanned for the presence of blood parasites using a light microscope Nikon Eclipse Ti - Arcturus XT of Applied Biosystems. Half of each blood smear was scanned at 400x magnification in search of larger parasites, including \u003cem\u003eLeucocytozoon\u003c/em\u003e, during 30\u0026ndash;45 min. The other half of each smear was scanned at 1000x magnification in search of \u003cem\u003eHaemoproteus\u003c/em\u003e, \u003cem\u003ePlasmodium\u003c/em\u003e and \u003cem\u003eLeucocytozoon\u003c/em\u003e parasites in up to \u0026gt;\u0026thinsp;10,000 erythrocytes per smear (see \u003csup\u003e\u003cspan citationid=\"CR34\" class=\"CitationRef\"\u003e34\u003c/span\u003e\u003c/sup\u003e) during 30 min. DNA from blood samples were extracted using the Maxwell\u0026reg;16 LEV system Research (Promega, Madison, WI) \u003csup\u003e\u003cspan citationid=\"CR35\" class=\"CitationRef\"\u003e35\u003c/span\u003e\u003c/sup\u003e. Samples were run in 0.8% agarose gels to check DNA integrity. To detect and identify avian parasites of the genera \u003cem\u003ePlasmodium\u003c/em\u003e, \u003cem\u003eHaemoproteus\u003c/em\u003e and \u003cem\u003eLeucocytozoon\u003c/em\u003e, we carried out nested PCRs following Hellgren et al. \u003csup\u003e\u003cspan citationid=\"CR36\" class=\"CitationRef\"\u003e36\u003c/span\u003e\u003c/sup\u003e to amplify a fragment of the mitochondrial cytochrome b gene. To check if the PCRs amplified successfully, we ran 3.0 \u0026micro;l of the final PCR product on a 2% agarose gel. We ran 6 repetitions of each PCR to exclude false negatives.\u003c/p\u003e \u003c/div\u003e"},{"header":"Declarations","content":"\u003cp\u003e\u003cstrong\u003eAcknowledgements\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eWe thank Karla Guerrero, Daniel Linke and Carrie Gunn for their assistance in the field, the Secretar\u0026iacute;a de Marina and fishermen from Nayarit for logistical support, and the SEMARNAT and CONANP for permission to work on Isla Isabel. Cristina Rodr\u0026iacute;guez provided logistical support, Gast\u0026oacute;n Contreras-Jim\u0026eacute;nez provided technical assistance in microscopy analysis, and Isabel Mart\u0026iacute;n and Cristina Perez provided technical assistance in the laboratory.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eAuthor contributions\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eSA and HD conceived the study. FRZ and SA sampled birds. FRZ, JMP, MJRL and JF analyzed the samples. SA drafted the manuscript with significant contributions of the other authors. All authors read and approved the manuscript.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eCompeting interests\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eThe authors declare no competing interests.\u003cstrong\u003e\u0026nbsp;\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eData availability\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eData supporting the conclusions are included within the article.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eEthics approval\u0026nbsp;\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eOur research complies with Mexican legal and ethical requirements. Permit to collect blood samples (SGPA/DGVS/01216617) was provided by Secretar\u0026iacute;a del Medio Ambiente y Recursos Naturales (SEMARNAT).\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eConsent for publication\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eNot applicable.\u0026nbsp;\u003c/p\u003e"},{"header":"References","content":"\u003col\u003e\u003cli\u003e\u003cspan\u003eAtkinson, C. T. \u0026amp; Van Riper, C. Pathogenicity and epizootiology of avian haematozoa: Plasmodium, Leucocytozoon, and Haemoproteus.\u003cem\u003eBird-parasite Interact.\u003c/em\u003e19\u0026ndash;48(1991).\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eSorci, G. \u0026amp; Moller, A. P. Comparative evidence for a positive correlation between haematozoan prevalence and mortality in waterfowl. \u003cem\u003eJ. Evol. Biol\u003c/em\u003e, \u003cb\u003e10\u003c/b\u003e, 731\u0026ndash;741 (1997).\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eMerino, S., Moreno, J., Sanz, J. J. \u0026amp; Arriero, E. Are avian blood parasites pathogenic in the wild? A medication experiment in blue tits (Parus caeruleus). Proc. R. Soc. B Biol. Sci. 267, 2507\u0026ndash;2510(2000).\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eAsghar, M. \u003cem\u003eet al.\u003c/em\u003e Hidden costs of infection: Chronic malaria accelerates telomere degradation and senescence in wild birds. \u003cem\u003eScience (80-.)\u003c/em\u003e, \u003cb\u003e347\u003c/b\u003e, 436\u0026ndash;438 (2015).\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eQuillfeldt, P., Arriero, E., Mart\u0026iacute;nez, J., Masello, J. F. \u0026amp; Merino, S. Prevalence of blood parasites in seabirds - a review. \u003cem\u003eFront. Zool\u003c/em\u003e, \u003cb\u003e8\u003c/b\u003e, 26 (2011).\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003ePiersma, T. Do Global Patterns of Habitat Use and Migration Strategies Co-Evolve with Relative Investments in Immunocompetence due to Spatial Variation in Parasite Pressure?, \u003cb\u003e80\u003c/b\u003e, 623 (1997).\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eMendes, L., Piersma, T., Lecoq, M., Spaans, B. \u0026amp; Ricklefs, R. E. Disease-limited distributions? Contrasts in the prevalence of avian malaria in shorebird species using marine and freshwater habitats., \u003cb\u003e109\u003c/b\u003e, 396\u0026ndash;404 (2005).\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eMart\u0026iacute;nez-Abra\u0026iacute;n, A., Esparza, B. \u0026amp; Oro, D. Lack of blood parasites in bird species: Does absence of blood parasite vectors explain it all? in Ardeola vol.\u0026nbsp;51 225\u0026ndash;232(2004).\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eCampioni, L. \u003cem\u003eet al.\u003c/em\u003e Absence of haemosporidian parasite infections in the long-lived Cory\u0026rsquo;s shearwater: evidence from molecular analyses and review of the literature. \u003cem\u003eParasitol. Res\u003c/em\u003e, \u003cb\u003e117\u003c/b\u003e, 323\u0026ndash;329 (2018).\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eOsorio-Beristain, M. \u0026amp; Drummond, H. Non-aggressive mate guarding by the blue-footed booby: A balance of female and male control. \u003cem\u003eBehav. Ecol. Sociobiol\u003c/em\u003e, \u003cb\u003e43\u003c/b\u003e, 307\u0026ndash;315 (1998).\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eNelson, J. B. \u003cem\u003ePelicans, cormorants and their relatives: The Pelecaniformes\u003c/em\u003e (Oxford University Press, 2006).\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eKim, S. Y., Torres, R., Dom\u0026iacute;nguez, C. A. \u0026amp; Drummond, H. Lifetime philopatry in the blue-footed booby: A longitudinal study. \u003cem\u003eBehav. Ecol\u003c/em\u003e, \u003cb\u003e18\u003c/b\u003e, 1132\u0026ndash;1138 (2007).\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eDrummond, H. \u0026amp; Rodr\u0026iacute;guez, C. Viability of booby offspring is maximized by having one young parent and one old parent. \u003cem\u003ePLoS One\u003c/em\u003e, \u003cb\u003e10\u003c/b\u003e, e0133213 (2015).\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eLee-Cruz, L. \u003cem\u003eet al.\u003c/em\u003e Prevalence of Haemoproteus sp. in Gal\u0026aacute;pagos blue-footed boobies: effects on health and reproduction.\u003cem\u003eParasitol. Open\u003c/em\u003e\u003cb\u003e2\u003c/b\u003e, (2016).\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eSantiago-Alarcon, D., Palinauskas, V. \u0026amp; Schaefer, H. M. Diptera vectors of avian Haemosporidian parasites: Untangling parasite life cycles and their taxonomy. \u003cem\u003eBiol. Rev\u003c/em\u003e, \u003cb\u003e87\u003c/b\u003e, 928\u0026ndash;964 (2012).\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eBond, J. G. \u003cem\u003eet al.\u003c/em\u003e Diversity of mosquitoes and the aquatic insects associated with their oviposition sites along the Pacific coast of Mexico. \u003cem\u003eParasites and Vectors\u003c/em\u003e, \u003cb\u003e7\u003c/b\u003e, 1\u0026ndash;19 (2014).\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eIba\u0026ntilde;ez-Bernal, S. Informe Final del proyecto Actualizaci\u0026oacute;n del Cat\u0026aacute;logo de Autoridad Taxon\u0026oacute;mica del orden Diptera (Insecta) de M\u0026eacute;xico CONABIO (JE006)(2017).\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eLevin, I. I. \u003cem\u003eet al.\u003c/em\u003e Hippoboscid-transmitted Haemoproteus parasites (Haemosporida) infect Galapagos Pelecaniform birds: Evidence from molecular and morphological studies, with a description of Haemoproteus iwa. \u003cem\u003eInt. J. Parasitol\u003c/em\u003e, \u003cb\u003e41\u003c/b\u003e, 1019\u0026ndash;1027 (2011).\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eMadsen, V. \u003cem\u003eet al.\u003c/em\u003e Testosterone levels and gular pouch coloration in courting magnificent frigatebird (Fregata magnificens): Variation with age-class, visited status and blood parasite infection. \u003cem\u003eHorm. Behav\u003c/em\u003e, \u003cb\u003e51\u003c/b\u003e, 156\u0026ndash;163 (2007).\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eClark, G. W. \u0026amp; Swinehart, B. Avian haematozoa from the offshore islands of northern Mexico. \u003cem\u003eWildl. Dis\u003c/em\u003e, \u003cb\u003e5\u003c/b\u003e, 111\u0026ndash;112 (1969).\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eQuillfeldt, P. \u003cem\u003eet al.\u003c/em\u003e Hemosporidian blood parasites in seabirds - A comparative genetic study of species from Antarctic to tropical habitats., \u003cb\u003e97\u003c/b\u003e, 809\u0026ndash;817 (2010).\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eMerino, S. \u003cem\u003eet al.\u003c/em\u003e Infection by haemoproteus parasites in four species of frigatebirds and the description of a new species of Haemoproteus (Haemosporida: Haemoproteidae). \u003cem\u003eJ. Parasitol\u003c/em\u003e, \u003cb\u003e98\u003c/b\u003e, 388\u0026ndash;397 (2012).\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eSvensson, L. M. E. \u0026amp; Ricklefs, R. E. Low diversity and high intra-island variation in prevalence of avian Haemoproteus parasites on Barbados, Lesser Antilles., \u003cb\u003e136\u003c/b\u003e, 1121\u0026ndash;1131 (2009).\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eLoiseau, C. \u003cem\u003eet al.\u003c/em\u003e Spatial variation of haemosporidian parasite infection in african rainforest bird species. \u003cem\u003eJ. Parasitol\u003c/em\u003e, \u003cb\u003e96\u003c/b\u003e, 21\u0026ndash;29 (2010).\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eMadsen, V. \u003cem\u003eFemale mate choice in the magnificent frigatebird (Fregata magnificens)\u003c/em\u003e (Universidad Nacional Aut\u0026oacute;noma de M\u0026eacute;xico, 2004).\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eSuper, P. E. \u0026amp; van Riper, C. A comparison of avian hematozoan epizootiology in two California coastal scrub communities. \u003cem\u003eJ. Wildl. Dis\u003c/em\u003e, \u003cb\u003e31\u003c/b\u003e, 447\u0026ndash;461 (1995).\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eCONANP. \u003cem\u003ePrograma de Conservaci\u0026oacute;n y Manejo del Parque Nacional Isla Isabel\u003c/em\u003e. (2005).\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eAncona, S., Drummond, H., Rodr\u0026iacute;guez, C. \u0026amp; Z\u0026uacute;\u0026ntilde;iga-Vega, J. J. Long-term population dynamics reveal that survival and recruitment of tropical boobies improve after a hurricane. \u003cem\u003eJ. Avian Biol\u003c/em\u003e, \u003cb\u003e48\u003c/b\u003e, 320\u0026ndash;332 (2017).\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eAncona, S., S\u0026aacute;nchez-Col\u0026oacute;n, S., Rodr\u0026iacute;guez, C. \u0026amp; Drummond, H. El Ni\u0026ntilde;o in the Warm Tropics: Local sea temperature predicts breeding parameters and growth of blue-footed boobies. \u003cem\u003eJ. Anim. Ecol\u003c/em\u003e, \u003cb\u003e80\u003c/b\u003e, 799\u0026ndash;808 (2011).\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eAnderson, D. J., Huyvaert, K. P. \u0026amp; Anchundia, D. Chronic lack of breeding by Gal\u0026aacute;pagos Blue-footed Boobies and associated population decline. Galapagos Report 2013\u003cem\u003e\u0026ndash;2014\u003c/em\u003e vol.\u0026nbsp;9 \u003cspan class=\"ExternalRef\"\u003e\u003cspan class=\"RefSource\"\u003ehttps://www.galapagos.org/wp-content/uploads/2015/09/GalapagosReport_2013-2014-22-Anderson-158-163.pdf\u003c/span\u003e\u003c/span\u003e (2015).\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eValkiunas, G. \u003cem\u003eet al.\u003c/em\u003e A comparative analysis of microscopy and PCR-based detection methods for blood parasites. \u003cem\u003eJ. Parasitol\u003c/em\u003e, \u003cb\u003e94\u003c/b\u003e, 1395\u0026ndash;1401 (2008).\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eSantiago-Alarcon, D. \u003cem\u003eet al.\u003c/em\u003e Parasites in space and time: a case study of haemosporidian spatiotemporal prevalence in urban birds. \u003cem\u003eInt. J. Parasitol\u003c/em\u003e, \u003cb\u003e49\u003c/b\u003e, 235\u0026ndash;246 (2019).\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eDrummond, H., Torres, R. \u0026amp; Krishnan, V. V. Buffered development: Resilience after aggressive subordination in infancy. \u003cem\u003eAm. Nat\u003c/em\u003e, \u003cb\u003e161\u003c/b\u003e, 794\u0026ndash;807 (2003).\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eMerino, S. \u0026amp; Potti, J. High Prevalence of Hematozoa in Nestlings of a Passerine Species, the Pied Flycatcher (Ficedula hypoleuca). \u003cem\u003eAuk\u003c/em\u003e, \u003cb\u003e112\u003c/b\u003e, 1041\u0026ndash;1043 (1995).\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eGuti\u0026eacute;rrez-L\u0026oacute;pez, R. \u003cem\u003eet al.\u003c/em\u003e Low prevalence of blood parasites in a long-distance migratory raptor: The importance of host habitat. \u003cem\u003eParasites and Vectors\u003c/em\u003e, \u003cb\u003e8\u003c/b\u003e, 1\u0026ndash;6 (2015).\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eHellgren, O., Waldenstr\u0026ouml;m, J. \u0026amp; Bensch, S. A new PCR assay for simultaneous studies of Leucocytozoon, Plasmodium, and Haemoproteus from avian blood. \u003cem\u003eJ. Parasitol\u003c/em\u003e, \u003cb\u003e90\u003c/b\u003e, 797\u0026ndash;802 (2004).\u003c/span\u003e\u003c/li\u003e\u003c/ol\u003e"}],"fulltextSource":"","fullText":"","funders":[],"hasAdminPriorityOnWorkflow":false,"hasManuscriptDocX":true,"hasOptedInToPreprint":true,"hasPassedJournalQc":"","hasAnyPriority":false,"hideJournal":true,"highlight":"","institution":"","isAcceptedByJournal":false,"isAuthorSuppliedPdf":false,"isDeskRejected":"","isHiddenFromSearch":false,"isInQc":false,"isInWorkflow":false,"isPdf":false,"isPdfUpToDate":true,"isWithdrawnOrRetracted":false,"journal":{"display":true,"email":"[email protected]","identity":"researchsquare","isNatureJournal":false,"hasQc":true,"allowDirectSubmit":true,"externalIdentity":"","sideBox":"","snPcode":"","submissionUrl":"/submission","title":"Research Square","twitterHandle":"researchsquare","acdcEnabled":true,"dfaEnabled":false,"editorialSystem":"","reportingPortfolio":"","inReviewEnabled":false,"inReviewRevisionsEnabled":true},"keywords":"Avian malaria, Plasmodium, Haemoproteus, Leucocytozoon, Wild birds, Seabirds","lastPublishedDoi":"10.21203/rs.3.rs-842608/v1","lastPublishedDoiUrl":"https://doi.org/10.21203/rs.3.rs-842608/v1","license":{"name":"CC BY 4.0","url":"https://creativecommons.org/licenses/by/4.0/"},"manuscriptAbstract":"\u003cp\u003eHaemosporidian parasites are common in birds, but often are not in seabirds. The absence of vectors/genetic resistance to infection have been proposed to explain this pattern. Examination of different host populations is required to confirm the absence of blood parasites in widespread host species, which could be differently exposed to blood parasites across their geographic range. Moreover, screening of blood parasites in many seabirds has been done only by visual inspection of blood smears, which can miss low-intensity infections. Screening of blood parasites of the genera \u003cem\u003ePlasmodium\u003c/em\u003e, \u003cem\u003eHaemoproteus\u003c/em\u003e and \u003cem\u003eLeucocytozoon\u003c/em\u003e, combining inspection of blood smears and PCR-based detection methods, revealed that a highly philopatric colony of blue-footed boobies (\u003cem\u003eSula nebouxii\u003c/em\u003e) in the Tropical North Pacific is likely free of these parasites. Earlier detection of \u003cem\u003eHaemoproteus\u003c/em\u003e parasites in frigatebirds cohabiting with boobies in our study site and blue-footed boobies breeding on the Galapagos Islands suggests that absence of blood parasites in this northern booby colony could not be attributable to the absence of vectors or genetic resistance to infection. High host specificity or fine-scale spatial heterogeneity in the abundance of insect vectors could explain our negative results, but these hypotheses remain to be tested. We emphasize the relevance of assessing the occurrence of blood parasites in different populations of widespread host species, such as blue-footed boobies.\u003c/p\u003e","manuscriptTitle":"Apparent Absence of Avian Malaria and Malaria-Like Parasites in Northern Blue-Footed Boobies Breeding On Isla Isabel","msid":"","msnumber":"","nonDraftVersions":[{"code":1,"date":"2021-08-27 13:42:26","doi":"10.21203/rs.3.rs-842608/v1","editorialEvents":[{"type":"communityComments","content":0}],"status":"published","journal":{"display":true,"email":"[email protected]","identity":"researchsquare","isNatureJournal":false,"hasQc":true,"allowDirectSubmit":true,"externalIdentity":"","sideBox":"","snPcode":"","submissionUrl":"/submission","title":"Research Square","twitterHandle":"researchsquare","acdcEnabled":true,"dfaEnabled":false,"editorialSystem":"","reportingPortfolio":"","inReviewEnabled":false,"inReviewRevisionsEnabled":true}}],"origin":"","ownerIdentity":"32c4acc4-3cd5-46b9-a0bd-15b924a90d6a","owner":[],"postedDate":"August 27th, 2021","published":true,"recentEditorialEvents":[],"rejectedJournal":[],"revision":"","amendment":"","status":"posted","subjectAreas":[{"id":6762790,"name":"Agroecology"},{"id":6762791,"name":"Epidemiology"}],"tags":[],"updatedAt":"2021-11-26T07:14:08+00:00","versionOfRecord":[],"versionCreatedAt":"2021-08-27 13:42:26","video":"","vorDoi":"","vorDoiUrl":"","workflowStages":[]},"version":"v1","identity":"rs-842608","journalConfig":"researchsquare"},"__N_SSP":true},"page":"/article/[identity]/[[...version]]","query":{"redirect":"/article/rs-842608","identity":"rs-842608","version":["v1"]},"buildId":"_2-kVJe1T_tPrBINL-cwx","isFallback":false,"isExperimentalCompile":false,"dynamicIds":[84888],"gssp":true,"scriptLoader":[]}