We studied the organization and temporal stability of an assemblage of malaria parasites (genera Plasmodium and Haemoproteus) and their passerine avian hosts in a forested study area in southern Missouri, USA, over four years. We detected parasite infections by polymerase chain reaction (PCR) of parasite DNA from host blood samples and identified parasite lineages by sequencing a part of the mitochondrial cytochrome b gene. We obtained 757 blood samples from 42 host species. Prevalence of malaria parasitism judged by PCR averaged 38.6% and varied in parallel in the three most abundant host species over the four years of the study. Parasite prevalence bore a U‐shaped relationship to host sample size. Prevalence was weakly positively associated with host body mass, but not with foraging stratum, nest height, nest type, plumage brightness, or sexual dichromatism. Over the sample as a whole, parasite prevalence did not vary between males and females or between hatch‐year and older individuals. We differentiated 34 parasite lineages. The number of host species per lineage varied from one to eight and increased with sample size. We recovered up to 14 lineages of parasite from a single host. Three relatively common lineages in the Ozarks were found nowhere else; four others were recovered from other sites in eastern North America; and six additional well‐sampled lineages were distributed in the Greater Antilles among resident island host species. Parasites that are endemic among native species of hosts on the tropical wintering grounds of Ozark birds were recovered from hatch‐year birds in the Ozarks, indicating that transmission takes place on the summer breeding grounds, and consequently, that suitable vectors are present in both the temperate and tropical portions of the parasite lineage distributions. We estimate that the number of parasite lineages within a local area will approximate the number of host species and that our perception of host breadth and parasite diversity will increase for most lineages and hosts with increased sampling. Thus, host–parasite relationships in a local area, including the role of parasites in sexual selection and the evolutionary maintenance of sex, are likely to be complex, with population and evolutionary dynamics involving many actors.

We describe a polymerase chain reaction (PCR) assay that detects avian malarial infection across divergent host species and parasite lineages representing both Plasmodium spp. and Haemoproteus spp. The assay is based on nucleotide primers designed to amplify a 286-bp fragment of ribosomal RNA (rRNA) coding sequence within the 6-kb mitochondrial DNA malaria genome. The rRNA malarial assay outperformed other published PCR diagnostic methods for detecting avian infections. Our data demonstrate that the assay is sensitive to as few as 10~5 infected erythrocytes in peripheral blood. Results of avian population surveys conducted with the rRNA assay suggest that prevalences of malarial infection are higher than previously documented, and that studies based on microscopic examination of blood smears may substantially underestimate the extent of parasitism by these apicomplexans. Nonetheless, because these and other published primers miss small numbers of infections detected by other methods, including inspection of smears, no assay now available for avian malaria is universally reliable.

The lark sparrow (Chondestes grammacus) is a ground-nesting passerine that breeds across much of the central North American steppe and sand barrens. Through genotyping and sequencing of avian malaria parasites we examined levels of malaria prevalence and determined the distribution of Haemoproteus and Plasmodium lineages across the breeding range of the lark sparrow. Analysis of 365 birds collected from five breeding locations revealed relatively high levels of malaria prevalence in adults (80 %) and juveniles (46 %), with infections being primarily of Haemoproteus (91 % of sequenced samples). Levels of genetic diversity and genetic structure of malaria parasites with respect to the avian host populations revealed distinct patterns for Haemoproteus and Plasmodium, most likely as a result of their distinct life histories, host specificity, and transmission vectors. With the exception of one common Haemoproteus haplotype detected in all populations, all other haplotypes were either population-specific or shared by two to three populations. A hierarchical analysis of molecular variance of Haemoproteus sequences revealed that 15-18 % of the genetic variation can be explained by differences among host populations/locations (p < 0.001). In contrast to the regional patterns of genetic differentiation detected for the lark sparrow populations, Haemoproteus parasites showed high levels of population-specific variation and no significant differences among regions, which suggests that the population dynamics of the parasites may be driven by evolutionary processes operating at small spatial scales (e.g., at the level of host populations). These results highlight the potential effects of host population structure on the demographic and evolutionary dynamics of parasites.