Diversity of avian blood parasites in wild passerines in Serbia with special reference to two new lineages

  • Daliborka StankovićEmail author
  • Jane Jönsson
  • Marko Raković
Original Article


Avian haemosporidians are vector-transmitted blood parasites that are distributed worldwide, abundant in many bird families and well studied across Europe and North America. Since avian haemosporidians have been poorly examined in the Palearctic migratory flyways of the Western Balkans, the goal of this study was to investigate which species of three haemosporidian genera, Plasmodium, Haemoproteus and Leucocytozoon, infect both resident and migratory passerines in Serbia. The prevalence, distribution and parasitaemia of avian haemosporidian infections were screened using both a nested PCR method and microscopy. Out of 202 birds sampled at seven localities, 66 were infected with haemosporidians, and the total prevalence was 32.7%. The great majority of infected birds (29 individuals) had moderate levels of parasitaemia. The most abundant haemosporidian genus was Haemoproteus at 26.1% prevalence. All infected birds were adults; none of the tested juveniles were infected. Mixed infection was only recorded in one bird. We identified 31 genetic lineages of haemosporidians. Two new cytochrome b lineages, of Plasmodium and Leucocytozoon, were identified and found in the hosts Common Chaffinch (Fringilla coelebs) and Golden Oriole (Oriolus oriolus), respectively. We identified three new host records for previously known lineages. The lineage GRW06 (Plasmodium elongatum) occurred in Common Chaffinch, while the lineages PARUS20 and PARUS25 (Leucocytozoon sp.) were recorded in Willow Tit (Poecile montanus) and Crested Tit (Lophophanes cristatus), respectively. We found statistically significant differences in the prevalence of three haemosporidian genera among residents and partial migrants. The difference in mean parasitaemia was significant only between residents and partial migrants.


Haemosporidian Passeriformes Plasmodium Haemoproteus Leucocytozoon 


Diversität der Blutparasiten bei wildlebenden Sperlingsvögeln in Serbien unter besonderer Beachtung zweier neuer Abstammungslinien

Vogel-Hämosporidien, weltweit verbreitete Blutparasiten, die durch Vektoren übertragen werden, kommen bei vielen Vogelfamilien häufig vor und sind in Europa und Nordamerika gut untersucht. Da über Vogel-Hämosporidien im paläarktischen Zugkorridor auf dem westlichen Balkan jedoch nur wenig bekannt ist, war das Ziel dieser Studie, herauszufinden, welche Arten der drei Hämosporidien-Gattungen Plasmodium, Haemoproteus und Leucocytozoon nicht-ziehende und ziehende Sperlingsvögel in Serbien infizieren. Die Prävalenz, die Verbreitung und die Parasitämie von Vogel-Hämosporidien-Infektionen wurden mit Hilfe von verschachtelter PCR und Mikroskopie ermittelt. Von 202 Vögeln, die an sieben Standorten beprobt wurden, waren 66 mit Hämosporidien infiziert. Die Gesamtprävalenz betrug 32,7%. Der Großteil der infizierten Vögel (29 Individuen) wies moderate Parasitämiespiegel auf. Die häufigste Hämosporidiengattung war Haemoproteus mit einer Prävalenz von 26,1%. Alle infizierten Tiere waren Altvögel, wohingegen keine der untersuchten Jungvögel infiziert waren. Eine gemischte Infektion wurde lediglich bei einem Vogel festgestellt. Wir haben 31 genetische Abstammungslinien von Hämosporidien-Parasiten identifiziert. Zwei neue Cytochrom b-Abstammungslinien von Plasmodium und Leucocytozoon wurden identifiziert und beim Buchfinken (Fringilla coelebs) und Pirolen (Oriolus oriolus) nachgewiesen. Zudem haben wir drei neue Wirte für zuvor bekannte Abstammungslinien identifiziert. Die Linie GRW06 (Plasmodium elongatum) kam bei Buchfinken vor, während die Linien PARUS20 und PARUS25 (Leucocytozoon sp.) bei Weidenmeisen (Poecile montanus) bzw. Haubenmeisen (Lophophanes cristatus) nachgewiesen wurden. Wir haben statistisch signifikante Unterschiede in der Prävalenz der drei Hämosporidiengattungen bei Standvögeln und Teilziehern gefunden. Der Unterschied in der mittleren Parasitämie war nur zwischen Standvögeln und Teilziehern signifikant.



The laboratory work was partially supported by the Natural History Museum in Belgrade, Serbia and the Department of Biology, Lund University, Sweden. Aleksandra Urošević is acknowledged for her help in the laboratory at the Institute for Medical Research, University of Belgrade, Serbia. We are thankful to Dr Staffan Bensch and two anonymous reviewers for valuable comments on earlier versions of the manuscript. We thank Dr Bojana Stanić for proofreading the manuscript and Nicola Crockford for correcting the English. The sampling in this study complies with the current legislation of the Ministry of Environmental Protection of the Republic of Serbia.

Supplementary material

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  1. Arizaga J, Esparza X, Barba E (2010) Haemosporidians in migratory Blackcaps (Sylvia atricapilla): a comparison between autumn and spring periods of passage. An de Biol 32:87–93Google Scholar
  2. Asghar M, Hasselquist D, Bensch S (2011) Are chronic avian haemosporidian infections costly in wild birds? J Avian Biol 42:530–537CrossRefGoogle Scholar
  3. Bennett GF, Peirce MA, Ashford RW (1993) Avian Haematozoa: mortality and pathogenicity. J Nat Hist 27(5):993–1001 (Taylor & Francis Online) CrossRefGoogle Scholar
  4. Bensch S, Åkesson S (2003) Temporal and spatial variation of hematozoans in Scandinavian Willow Warblers. J Parasitol 89:388–391CrossRefGoogle Scholar
  5. Bensch S, Stjernman M, Hasselquist D, Östman Ö, Hansson B, Westerdahl H, Pinheiro RT (2000) Host specificity in avian blood parasites: a study of Plasmodium and Haemoproteus mitochondrial DNA amplified from birds. Proc R Soc Lond B 267:1583–1589CrossRefGoogle Scholar
  6. Bensch S, Waldenström Ј, Jonzén N, Westerdahl H, Hansson B, Sejberg D, Hasselquist D (2007) Temporal dynamic and diversity of avian malaria parasites in a single host species. J Anim Ecol 76:112–122CrossRefGoogle Scholar
  7. 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 9:1353–1358CrossRefGoogle Scholar
  8. Benson DA, Cavanaugh M, Clark K, Karsch-Mizrachi I, Lipman DJ, Ostell J, Sayers EW (2014) GenBank. Nucleic Acids Res 41:36–42CrossRefGoogle Scholar
  9. Bentz S, Rigaud T, Barroca M, Martin-Laurent F, Bru D, Moreau J, Faivre B (2006) Sensitive measure of prevalence and parasitaemia of haemosporidia from European Blackbird (Turdus merula) populations: value of PCR-RFLP and quantitative PCR. Parasitology 133:685–692CrossRefGoogle Scholar
  10. Berthold P (1996) Control of bird migration. Chapman and Hall, London, pp 25–26Google Scholar
  11. Cramp S (ed) (1988) The birds of the Western Palearctic, vol V. Oxford University PressGoogle Scholar
  12. del Hoyo J, Collar NJ (2014) HBW and BirdLife International illustrated checklist of the birds of the world: passerines, vol 2. Lynx, BarcelonaGoogle Scholar
  13. Diarra M, Fall M, Fall GA, Diop A, Seck MT, Garros C, Balenghien T, Allène X, Rakotoarivony I, Lancelot R, Mall I, Bakhoum M, Dosum AM, Ndao M, Bouyer J, Guis H (2014) Seasonal dynamics of Culicoides (Diptera: Ceratopogonidae) biting midges, potential vectors of African horse sickness and bluetongue viruses in the Niayes area of Senegal. Parasites Vectors 7:147CrossRefGoogle Scholar
  14. Dimitrov D, Zehtindjiev P, Bensch S (2010) Genetic diversity of avian blood parasites in SE Europe: cytochrome b lineages of the genera Plasmodium and Haemoproteus (Haemosporida) from Bulgaria. Acta Parasitol 55:201–209CrossRefGoogle Scholar
  15. Fallon SM, Ricklefs RE, Swanson BL, Bermingham E (2003) Detecting avian malaria: an improved polymerase chain reaction diagnostic. J Parasitol 89(5):1044–1047CrossRefGoogle Scholar
  16. Fallon SM, Fleischer RC, Graves GR (2006) Malarial parasites as geographical markers in migratory birds? Biol Let 2:213–216CrossRefGoogle Scholar
  17. Godfrey RD, Fedynich AM, Pence DB (1987) Quantification of hematozoa in blood smears. J Wildl Dis 23:558–565CrossRefGoogle Scholar
  18. Hall TA (1999) BIOEDIT: a user friendly biological sequence alignment editor and analysis program for Windows 95/98 NT. Nucleic Acid Symp Ser 41:95–98Google Scholar
  19. Hatchwell BJ, Wood MJ, Anwar M, Perrins CM (2000) The prevalence and ecology of the haematozoan parasites of European Blackbirds, Turdus merula. Can J Zool 78(4):684–687CrossRefGoogle Scholar
  20. 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(4):797–802CrossRefGoogle Scholar
  21. Hellgren O, Pérez-Tris J, Bensch S (2009) A jack-of-all-trades and still a master of some: prevalence and host range in avian malaria and related blood parasites. Ecology 90(10):2840–2849CrossRefGoogle Scholar
  22. Hellgren O, Križanauskiené A, Hasselquist D, Bensch S (2011) Low haemosporidian diversity and one key-host species in a bird malaria community on a mid-Atlantic island (São Miguel, Azores). J Wildl Dis 47(4):849–859CrossRefGoogle Scholar
  23. Latta SC, Ricklefs RE (2010) Prevalence patterns of avian haemosporida on Hispaniola. J Avian Biol 41:25–33CrossRefGoogle Scholar
  24. MalAvi. A database for avian haemosporidian parasites. Accessed 23 Dec 2017
  25. Palinauskas V, Markovets MY, Kosarev VV, Efremov VD, Sokolov LV, Valkiûnas G (2005) Occurrence of avian haematozoa in Ekaterinburg and Irkutsk districts of Russia. Ekologija (Lithuania) 4:8–12Google Scholar
  26. Pérez-Tris J, Bensch S (2005) Diagnosing genetically diverse avian malarial infections using mixed-sequence analysis and TA-cloning. Parasitology 131(1):15–23CrossRefGoogle Scholar
  27. R Core Team (2016) R: a language and environment for statistical computing. R Foundation for Statistical Computing, Vienna. Accessed 14 Oct 2017
  28. Richardson DS, Jury FL, Blaakmeer J, Komdeur J, Burke T (2001) Parentage assignment and extra—group paternity in a cooperative breeder: the Seychelles Warbler (Acrocephalus sechellensis). Mol Ecol 10:2263–2273CrossRefGoogle Scholar
  29. Ricklefs RE, Swanson BL, Fallon SM, Martínez-Abraín L, Cheuerlein A, Gray J, Latta SC (2005) Community relationships of avian malaria parasites in southern Missouri. Ecol Monogr 75(4):543–559CrossRefGoogle Scholar
  30. Rönn JAC, Harrod C, Bensch S, Wolf JBW (2015) Transcontinental migratory connectivity predicts parasite prevalence in breeding populations of the European Barn Swallow. J Evol Biol 28:535–546CrossRefGoogle Scholar
  31. Scheuerlein A, Ricklefs R (2004) Prevalence of blood parasites in European passeriform birds. Proc R Soc Lond B 271:1363–1370CrossRefGoogle Scholar
  32. Schmid S, Fachet K, Dinkel A, Mackenstedt U, Woog F (2017) Carrion Crows (Corvus corone) of southwest Germany: important hosts for haemosporidian parasites. Malar J 16(1):369CrossRefGoogle Scholar
  33. Shurulinkov P, Golemansky V (2003) Plasmodium and Leucocytozoon (Sporozoa: Haemosporida) of wild birds in Bulgaria. Acta Protozool 42(3):205–214Google Scholar
  34. Sibley CG, Monroe BL (1990) Distribution and taxonomy of birds of the world. Yale University Press, New Haven, LondonGoogle Scholar
  35. Sorensen MC, Asghar M, Bensch S, Fairhurst GD, Jenni-Eiermann S, Spottiswoode CN (2016) A rare study from the wintering ground provides insight into the cost of malaria infection for migratory birds. J Avian Biol 47:575–582CrossRefGoogle Scholar
  36. Svensson L (1992) Identification guide to European passerines, 4th edn. British Trust for Ornithology, StockholmGoogle Scholar
  37. Tamura K, Stecher G, Peterson D, Filipski A, Kumar S (2013) MEGA6: molecular evolutionary genetics analysis version 6.0. Mol Biol Evol 30:2725–2729CrossRefGoogle Scholar
  38. Valkiūnas G (2005) Avian malaria parasites and other haemosporidia. CRC, Boca RatonGoogle Scholar
  39. Valkiūnas G, Iezhova T, Golemansky G, Pilarska D, Zehtindjiev P (1999) Blood protozoan parasites (Protozoa: Kinetoplastida and Haemosporida) in wild birds from Bulgaria. Acta Zool Bulg 51:127–129Google Scholar
  40. Valkiūnas G, Iezhova TA, Križanauskienė A, Palinauskas V, Sehgal RNM (2008a) A comparative analysis of microscopy and PCR-based detection methods for blood parasites. J Parasitol 94(6):1395–1401CrossRefGoogle Scholar
  41. Valkiūnas G, Zehtindjiev P, Dimitrov D, Križanauskienė A, Iezhova TA, Bensch S (2008b) Polymerase chain reaction-based identification of Plasmodium (Huffia) elongatum, with remarks on species identity of haemosporidian lineages deposited in GenBank. Parasitol Res 102:1185–1193CrossRefGoogle Scholar
  42. Ventim R, Tenreiro P, Grade N, Encarnação P, Araújo M, Mendes L, Pérez-Tris J, Ramos JA (2012) Characterization of haemosporidian infections in warblers and sparrows at south-western European reed beds. J Ornithol 153:505–512CrossRefGoogle Scholar
  43. Waldenström J, Bensch S, Kiboi S, Hasselquist D, Ottosson U (2002) Cross-species infection of blood parasites between resident and migratory songbirds in Africa. Mol Ecol 11:1545–1554CrossRefGoogle Scholar
  44. Wiersch SC, Lubjuhn T, Maier WA, Kampen H (2007) Haemosporidian infection in passerine birds from Lower Saxony. J Ornithol 148:17–24CrossRefGoogle Scholar
  45. Zehtindjiev P, Ilieva M, Križanauskienė A, Oparina O, Oparin M, Bensch S (2009) Occurrence of haemosporidian parasites in the Paddyfield Warbler, Acrocephalus agricola (Passeriformes, Sylviidae). Acta Parasitol 54(4):295–300CrossRefGoogle Scholar
  46. Zehtindjiev P, Križanauskienė A, Bensch S, Palinauskas V, Asghar M, Dimitrov D, Scebba S, Valkiūnas G (2012) A new morphologically distinct avian malaria parasite that fails detection by established polymerase chain reaction-based protocols for amplification of the cytochrome b gene. J Parasitol 98(3):657–665CrossRefGoogle Scholar
  47. Zuk M (1990) Reproductive strategies and diseases susceptibility, an evolutionary viewpoint. Parasitol Today 6(7):231–233CrossRefGoogle Scholar

Copyright information

© Deutsche Ornithologen-Gesellschaft e.V. 2019

Authors and Affiliations

  • Daliborka Stanković
    • 1
    Email author
  • Jane Jönsson
    • 2
  • Marko Raković
    • 1
  1. 1.Natural History MuseumBelgradeRepublic of Serbia
  2. 2.Department of Biology, Ecology BuildingLund UniversityLundSweden

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