Haemosporida parasites of even-toed ungulates are diverse and globally distributed, but since their discovery in 1913 their characterization has relied exclusively on microscopy-based descriptions. In order to bring molecular approaches to bear on the identity and evolutionary relationships of ungulate malaria parasites, we conducted Plasmodium cytb-specific nested PCR surveys using blood from water buffalo in Vietnam and Thailand, and goats in Zambia. We found that Plasmodium is readily detectable from water buffalo in these countries, indicating that buffalo Plasmodium is distributed in a wider region than India, which is the only area in which buffalo Plasmodium has been reported. Two types (I and II) of Plasmodium sequences were identified from water buffalo and a third type (III) was isolated from goat. Morphology of the parasite was confirmed in Giemsa-reagent stained blood smears for the Type I sample. Complete mitochondrial DNA sequences were isolated and used to infer a phylogeny in which ungulate malaria parasites form a monophyletic clade within the Haemosporida, and branch prior to the clade containing bird, lizard and other mammalian Plasmodium. Thus it is likely that host switching of Plasmodium from birds to mammals occurred multiple times, with a switch to ungulates independently from other mammalian Plasmodium.

Plasmodium was first identified in a goat in Angola in 1923, and only recently characterized by DNA isolation from a goat blood sample in Zambia. Goats were first domesticated in the Fertile Crescent approximately 10,000 years ago, and are now globally distributed. It is not known if the Plasmodium identified in African goats originated from parasites circulating in the local ungulates, or if it co-evolved in the goat before its domestication. To address this question, we performed PCR-based surveillance using a total of 1,299 goat blood samples collected from Sudan and Kenya in Africa, Iran in west Asia, and Myanmar and Thailand in southeast Asia. Plasmodium DNA was detected from all locations, suggesting that the parasite is not limited to Africa, but widely distributed. Whole mitochondrial DNA sequences revealed that there was only one nucleotide substitution between Zambian/Kenyan samples and others, supporting the existence of a goat-specific Plasmodium species, presumably Plasmodium caprae, rather than infection of goats by local ungulate malaria parasites. We also present the first photographic images of P. caprae, from one Kenyan goat sample.

Characterising the genomic variation and population dynamics of Plasmodium falciparum parasites in high transmission regions of Sub-Saharan Africa is crucial to the long-term efficacy of regional malaria elimination campaigns and eradication. Whole-genome sequencing (WGS) technologies can contribute towards understanding the epidemiology and structural variation landscape of P. falciparum populations, including those within the Lake Victoria basin, a region of intense transmission. Here we provide a baseline assessment of the genomic diversity of P. falciparum isolates in the Lake region of Kenya, which has sparse genetic data. Lake region isolates are placed within the context of African-wide populations using Illumina WGS data and population genomic analyses. Our analysis revealed that P. falciparum isolates from Lake Victoria form a cluster within the East African parasite population. These isolates also appear to have distinct ancestral origins, containing genome-wide signatures from both Central and East African lineages. Known drug resistance biomarkers were observed at similar frequencies to those of East African parasite populations, including the S160N/T mutation in the pfap2mu gene, which has been associated with delayed clearance by artemisinin-based combination therapy. Overall, our work provides a first assessment of P. falciparum genetic diversity within the Lake Victoria basin, a region targeting malaria elimination.

We report, for the first time, the presence of ungulate malaria parasites in South America. We conducted PCR-based surveys of blood samples of multiple deer species and water buffalo from Brazil and detected Plasmodium sequences from pampas deer (Ozotoceros bezoarticus) samples. Phylogenic analysis revealed that the obtained sequences are closely related to the Plasmodium odocoilei clade 2 sequence from North American white-tailed deer (Odocoileus virginianus). Nucleotide differences suggest that malaria parasites in South American pampas deer and North American P. odocoilei clade 2 branched more recently than the Great American Interchange.

Several vector-borne pathogens restrict livestock farming and have significant economic impact worldwide. In endemic areas livestock are exposed to different tick species carrying various pathogens which could result in co-infection with several tick-borne pathogens in a single host. Although the co-infection of and the interaction among pathogens are critical factors to determine the disease outcome, pathogen interactions in the vector and the host are poorly understood. In this study, we surveyed the presence of Babesia ovis , Theileria ovis , Theileria lestoquardi , Anaplasma ovis , Anaplasma phagocytophilum , and Anaplasma marginale in 200 goats from 3 different districts in Sistan and Baluchestan province, Iran. Species-specific diagnostic PCRs and sequence analysis revealed that 1.5%, 12.5%, and 80% of samples were positive for T . lestoquardi , T . ovis , and A . ovis , respectively. Co-infections of goats with up to 3 pathogens were seen in 22% of the samples. We detected a significant association between T . ovis infection and age, T . ovis infection and location (Zabol), and A . ovis infection and location (Sarbaz) by multivariate logistic regression analysis. In addition, by analyzing the data with respect to Plasmodium caprae infection in these goats, a negative correlation was found between P . caprae and A . ovis infection. This study contributes to understanding the epidemiology of vector-borne pathogens and their interplay in goats.

Background Plasmodium falciparum SURFIN4.1 is a putative ligand expressed on the merozoite and likely on the infected red blood cell, whose gene was suggested to be under directional selection in the eastern Kenyan population, but under balancing selection in the Thai population. To understand this difference, surf 4.1 sequences of western Kenyan P. falciparum isolates were analysed. Frameshift mutations and copy number variation (CNV) were also examined for the parasites from western Kenya and Thailand.ResultsPositively significant departures from neutral expectations were detected on the surf 4.1 region encoding C-terminus of the variable region 2 (Var2) by 3 population-based tests in the western Kenyan population as similar in the Thai population, which was not covered by the previous analysis for eastern Kenyan population. Significant excess of non-synonymous substitutions per nonsynonymous site over synonymous substitutions per synonymous site was also detected in the Var2 region. Negatively significant departures from neutral expectations was detected on the region encoding Var1 C-terminus consistent to the previous observation in the eastern Kenyan population. Parasites possessing a frameshift mutation resulting a product without intracellular Trp-rich (WR) domains were 22/23 in western Kenya and 22/36 in Thailand. More than one copy of surf 4.1 gene was detected in western Kenya (4/24), but no CNV was found in Thailand (0/36).ConclusionsThe authors infer that the high polymorphism of SURFIN4.1 Var2 C-terminus in both Kenyan and Thai populations were shaped-up by diversifying selection and maintained by balancing selection. These phenomena were most likely driven by immunological pressure. Whereas the SURFIN4.1 Var1 C-terminus is suggested to be under directional selection consistent to the previous report for the eastern Kenyan population. Most western Kenyan isolates possess a frameshift mutation that would limit the expression of SURFIN4.1 on the merozoite, but only 60% of Thai isolates possess this frameshift, which would affect the level and type of the selection pressure against this protein as seen in the two extremities of Tajima’s D values for Var1 C-terminus between Kenyan and Thai populations. CNV observed in Kenyan isolates may be a consequence of this frameshift mutation to increase benefits on the merozoite surface.

Theileria, Babesia, and Anaplasma are tick-borne pathogens affecting livestock industries worldwide. In this study, we surveyed the presence of Babesia ovis, Theileria ovis, Theileria lestoquardi, Anaplasma ovis,Anaplasma phagocytophilum, and Anaplasma marginale in 200 goats from 3 different districts in Sistan and Baluchestan province, Iran. Species-specific diagnostic PCR and sequence analysis revealed that 1.5%, 12.5%, and 80% of samples were positive for T. lestoquardi, T. ovis, and A. ovis, respectively. Co-infections of goats with up to 3 pathogens were seen in 22% of the samples. We observed a positive correlation between A. ovis and T. ovis infection. In addition, by analyzing the data with respect to Plasmodium caprae infection in these goats, a negative correlation was found between P. caprae and A. ovis and between P. caprae and T. ovis. This study contributes to understanding the epidemiology of vector-borne pathogens and their interplay in goats.

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