Molecular and statistical approaches to the delimitation of Eimeriidae species: a case of extreme polymorphism in eimerian oocysts from the plumbeous pigeon Patagioenas plumbea (Vieillot, 1818) (Columbiformes) in South America.

The current work aimed to analyze, morphologically, statistically, and molecularly, oocysts shed from plumbeous pigeons, Patagioenas plumbea (Vieillot, 1818), from a locality at 2197 m of altitude near the Agulhas Negras peak, the highest point of the State of Rio de Janeiro, southeastern Brazil. The oocysts were extremely polymorphic, being subspheroidal, ovoidal, or ellipsoidal, in addition to having the random presence/absence of characteristic features associated with the oocyst wall, such as micropyle, micropyle cap, lateral micropyle, and outer veil/rough wall. Linear regression confirmed the extreme polymorphism of oocysts, showing that if all combinations of taxonomic characters in oocysts (morphotypes) were overestimated, 19 different species could be identified/described. In contrast, the means comparison analysis between oocysts with the presence/absence of characteristic features and the histograms showed equivalences and regularity in the distribution in the classes of measures, which indicate the presence of a single species in the measured oocysts. Molecular analyses were performed from the isolation of individual oocysts of different morphotypes, which had their genetic material extracted, amplified, and sequenced in 4 non-overlapping loci in the cox1 and cox3 genes and fragments of the small and large subunit rDNA of mitochondrial DNA. The sequences were 100% identical between the morphotypes, with the exception of a very small divergence observed at the locus that partially covers the cox3 gene. The phylogenetic analysis was inconclusive for the locus within the cox1 gene traditionally used for eimeriid coccidians; however, the other loci should have a promising future for phylogenetic studies when more sequences for the same genic regions are deposited in GenBank. Finally, the multifactorial analysis of the current work supported that the polymorphic oocysts shed from P. plumbea are a single species, which was named Eimeria patagioenasae, making this the twenty-second eimerian description from Columbiformes.

Isospora basileuterusi n. sp. (Apicomplexa: Eimeriidae) from the golden-crowned warbler Basileuterus culicivorus (Deppe) (Passeriformes: Parulidae) in South America.

Isospora basileuterusi Mello & Berto n. sp. is described based on material from the golden-crowned warbler Basileuterus culicivorus (Deppe) captured in the Itatiaia National Park (Parque Nacional do Itatiaia), a conservation unit in south-eastern Brazil. Oöcysts of the new species are ellipsoidal to ovoidal, measuring on average 25.2 × 21.1 µm, with a smooth, bi-layered wall, c.1.6 µm thick. Micropyle and oöcyst residuum are both absent, but one to three polar granules are present. Sporocysts are ellipsoidal to lemon-shaped, measuring on average 15.3 × 9.5 µm, with a knob-like Stieda body and a trapezoidal sub-Stieda body. Sporocyst residuum is present, usually as a body of membrane-bound granules. Sporozoites are vermiform, with refractile bodies. Four of the 19 warblers captured (21%) were infected with the new species. Molecular analysis of the mitochondrial cytochrome c oxidase subunit 1 (cox1) gene revealed a similarity of 99.5% between the new species and Isospora serinuse Yang, Brice, Elliot & Ryan, 2015 from island canaries Serinus canaria (L.) in Western Australia. The oöcysts of I. basileuterusi n. sp. can be distinguished from the four other Isospora spp. recorded in hosts of the Parulidae, and from the molecularly most closely related species, by the typical ellipsoidal to lemon-shaped sporocysts, with small sub-Stieda body and a membrane-bound sporocyst residuum. Therefore, based on the morphological and molecular features, I. basileuterusi n. sp. is the fifth species described in a host of the family Parulidae and the first molecularly characterized via sequencing the cox1 gene.

Molecular identification of Isospora coerebae Berto, Flausino, Luz, Ferreira amp; Lopes, 2010 (Chromista: Miozoa: Eimeriidae) from the bananaquit Coereba flaveola (Linnaeus, 1758) (Passeriformes: Thraupidae: Coerebinae) from Brazil.

Isospora coerebae Berto, Flausino, Luz, Ferreira Lopes, 2010 is a coccidian protozoan described from bananaquits Coereba flaveola (Linnaeus, 1758), on Marambaia Island, which is located on the southeastern Brazilian coast. In this current work, I. coerebae is identified from C. flaveola in a protected area close to Marambaia Island, but on the mainland, establishing a new location of parasitism, in addition to providing a preliminary genotypic characterization via sequencing of two regions of the mitochondrial cytochrome c oxidase subunit 1 (COI) gene. Its oocysts are sub-spherical, 24.4 22.9 m, with smooth, bilayered wall, ~1.7 m thick. Micropyle, polar granules and oocyst residuum are absent. Sporocysts are elongate ovoidal, 17.6 10.5 m. Stieda body prominent and rounded and sub-Stieda body short and wide. Sporocyst residuum is composed of scattered granules of different sizes. Sporozoites are vermiform with a prominent posterior refractile body. The oocysts of the current work are morphologically equivalent to the original description of I. coerebae, which have a typical and easily identifiable morphology, mainly in the Stieda and Sub-Stieda bodies. The two sequenced gene regions of the COI gene approximated I. coerebae to Isospora spp. from Southeastern Brazil, but also from Isospora spp. from passerines of North America, Europe and Asia. Although there is a small overlap between the two genic regions sequenced in the current work, it is estimated that the longer COI sequence, which was recently designed and still not widespread, should show better phylogenetic results in the future.

Isospora brasilsatoae n. sp. (Apicomplexa: Eimeriidae) from thrushes Turdus spp. (Passeriformes: Turdidae) from Brazil.

A new species of coccidia (Protozoa: Apicomplexa: Eimeriidae) found parasitizing the yellow-legged thrush Turdus flavipes (Vieillot, 1818), the pale-breasted thrush Turdus leucomelas Vieillot, 1818 and the creamy-bellied thrush Turdus amaurochalinus Cabanis, 1850 in Southeastern Brazil is described. Isospora brasilsatoae n. sp. has oocysts that are ovoidal to pyriform, 26.8 × 21.1 µm, with smooth, bilayered wall, ~1.6 µm thick. Micropyle present. Oocyst residuum absent, but one to three polar granules are present. Sporocysts are elongate ovoidal, 18.1 × 10.5 µm. Stieda body knob-like and sub-Stieda body trapezoidal. Sporocyst residuum is composed of granules that appear to be membrane-bounded. Sporozoites are vermiform with refractile bodies and striations. This is the ninth description of an Isospora sp. parasitizing a New World thrush.