Haematozoa

Smith, Matthew M. & Ramey, Andrew M., 2015, Prevalence and genetic diversity of haematozoa in South American waterfowl and evidence for intercontinental redistribution of parasites by migratory birds, International Journal for Parasitology: Parasites and Wildlife 4 (1), pp. 22-28 : 24-25

3.1. Haematozoa detection and prevalence

Of the 804 waterfowl blood samples screened for haematozoa infection, five samples failed to amplify the region of the COI gene used as a positive control. These samples were dropped from further analyses since we were unable to verify the presence of DNA in each extraction. Of the remaining 799 samples, 25 were positive for haematozoa infection (3.1% prevalence). Two samples were coinfected with parasites of multiple genera for a total of 27 haematozoa infections ( Table 1 ). Eight, five, and fourteen samples were identified as positive for Leucocytozoon , Haemoproteus , and Plasmodium parasites, respectively ( Table 1 ). Of the 25 parasitepositive blood samples, 24 came from hosts collected at field sites in Peru, with only a single positive sample originating from Argentina ( Table 1 ). Infections by haematozoan parasites were confined to only six of eleven waterfowl species screened, with the majority of infections identified in Torrent Ducks ( Merganetta armata ; Table 1 ).

3.2. Haplotype diversity

Analysis of haematozoa mtDNA cytochrome b sequences revealed a total of twelve unique haplotypes among South American waterfowl samples ( Table 2 ; Fig. 2 ). Haplotypes were identified as Plasmodium (n = 5), Leucocytozoon (n = 4), and Haemoproteus (n = 3). Only three of the twelve haplotypes detected in our samples occurred in multiple individuals (Haem 1, Plas 3, and Leuc 1; Fig. 2 ). Haemoproteus haplotype Haem 1 was identified from two birds of separate species, while Plasmodium haplotype Plas 3 was found in six different individuals, all of which were Torrent Ducks. Leucocytozoon haplotype Leuc 1 was observed in five different individuals of four species ( Table 2 ).

Eight haplotypes detected in our samples from South American waterfowl were identical to parasite lineages reported on the MalAvi and GenBank databases. Leucocytozoon haplotypes Leuc 1 and Leuc 4 were identical to lineages TUSW04 and TUSW05 respectively, which were detected in Alaska Tundra Swans and California Northern Pintails ( Ramey et al., 2012, 2013; Table 2 ). South American haplotype Leuc 2 was identical to lineage NOPI04 detected in Northern Pintails ( Anas acuta ) in California ( Ramey et al., 2013). Haemoproteus haplotype Haem 1 was identical to CYGNUS01, which has been previously identified in Tundra Swans in Minnesota and Alaska, Northern Pintails in California, as well as a Mallard ( Anas platyrhynchos ) from Minnesota ( Ricklefs and Fallon, 2002; Ramey et al., 2012, 2013). Four Plasmodium mtDNA cytochrome b haplotypes from our samples, Plas 2, Plas 3, Plas 4, and Plas 5, were identical to MILANS05, TFUS05, PESA01, and PADOM11 lineages, respectively. MILANS05 was documented infecting Black Kites ( Milvus migrans ) in Europe ( Pérez-Rodríguez et al., 2013), as well as Ruffs ( Philomachus pugnax ) in Africa ( Mendes et al., 2013). Lineage TFUS05 was detected infecting Great Thrushes ( Turdus fuscater ) in South America ( Lotta et al., 2013). PESA01 was observed in three separate host species: a Pectoral Sandpiper ( Calidris melanotos ) on the Arctic coast of Alaska ( Yohannes et al., 2008), a White-tipped Dove ( Leptotila verreauxi ) in Uruguay, and a Mouse-colored Tyrannulet ( Phaeomyias murina ) in Brazil ( Durrant et al., 2006; Lacorte et al., 2013). Lastly, the lineage PADOM11 is a common lineage of Plasmodium , documented infecting over 20 host species in North America, South America, and Asia (e.g. Durrant et al., 2006; Martinsen et al., 2006).

3.3. Phylogenetic analysis

Phylogenetic analysis supported structuring of avian parasite mtDNA haplotypes by genera. For Haemoproteus and Leucocytozoon parasites, phylogenetic analysis provided strong support for clades comprised of lineages originating from waterfowl sampled in both North and South America, and other clades comprised entirely of parasite mtDNA sequences originating from passerines sampled in South America ( Fig. 3 ). Sub-structuring of Haemoproteus and Leucocytozoon clades by continent of origin was not well supported for lineages detected in waterfowl. Avian Plasmodium lineages clustered into two groups of closely related sequences. Both groups of closely related Plasmodium lineages contained sequences originating from both North America and South America.

Figures

Fig. 2. Minimum spanning network for haematozoa mitochondrial DNA cytochrome b haplotypes detected in South American waterfowl. Shaded circles represent unsampled nodes. All circles are drawn proportional to the frequency at which haplotypes were observed. Lines separating nodes are drawn to scale based on the number of nucleotide mutations, unless otherwise indicated by hash marks. Only haplotypes with a length of 358 bp or greater were included. Haplotype name abbreviations are as follows: Haem = Haemoproteus, Leuc = Leucocytozoon, and Plas = Plasmodium.

Fig. 3. Bayesian phylogenetic tree of haematozoa mitochondrial DNA cytochrome b haplotypes obtained from infected waterfowl. Trees were rooted with mammalian Plasmodium outgroups. Node tips are labeled with parasite genus (Haem = Haemoproteus, Leuc = Leucocytozoon, and Plas = Plasmodium), followed by the lineage name, GenBank accession number for each sequence, host order (passerine/waterfowl), and the country/state from which the samples were collected. All haplotypes identified in this study are highlighted in red. Numbers on branches represent posterior probabilities from the analysis. Asterisks after node tip labels indicate sequences from our study that were identical to lineages previously found in non-waterfowl hosts. All reference sequences were obtained from the National Center for Biotechnology Information website.