Molecular Screening of Haemogregarine Hemoparasites (Apicomplexa: Adeleorina: Haemogregarinidae) in Populations of Native and Introduced Pond Turtles in Eastern Europe

Abstract

Haemogregarines (Apicomplexa: Adeleorina) are the most common and widespread reptilian blood parasites. Haemogregarina stepanowi was the first haemogregarine described from a reptile, the European pond turtle Emys orbicularis, and initial assessments indicated it was widespread across different pond turtle host species across much of Europe, the Middle East and North Africa. However, recent molecular assessments have indicated the presence of multiple genetically distinct forms in North Africa and the Iberian Peninsula, and extensive mixed infections which may be associated with a negative impact on the hosts. Here, we screened two native species, E. orbicularis and Mauremys rivulata, and the introduced Trachemys scripta from Serbia and North Macedonia for haemogregarines by amplifying and sequencing part of the 18S rRNA gene of these parasites, and used a standard DNA barcoding approach to identify leeches, the final host, attached to pond turtles. Our results again demonstrate the occurrence of considerable diversity of parasites in the analysed pond turtle species, and that T. scripta are likely infected by local haemogregarine parasites, and not those that are found in its native range. Leeches were identified as Placobdella costata, part of a lineage from Northern Europe. Mixed infections within pond turtles were again common. Current haemogregarine taxonomy does not reflect the genetic diversity identified, and a full taxonomic reassessment is needed.

Molecular Phylogenies of Leeches and Haemoparasites Infecting Freshwater Turtles in Aquatic Ecosystems of Northern Africa Suggest Phylogenetic Congruence between Placobdella costata Sensu Lato and Haemogregarina stepanowi Sensu Lato

“…While levels of COI genetic variations clearly indicate the occurrence of several haemogregarine species within the same host species in sympatry, even in syntopy, which is illustrated by numerous cases of coinfections, it is still extremely difficult to link a particular molecular lineage to a specific morphological species, as also reported in [ 17 ]. While the first molecular investigations on Haemogregarina have shown from nuclear markers that H. stepanowi possesses a wide distribution though Europe, Turkey, and the Middle East to Iran with a very low host-specificity [ 12 ], the most recent advents in molecular systematics of H. stepanowi have shown that this species could be actually split into a great number of distinct cryptic haemogregarine species [ 15 , 19 , 52 , 53 ], infecting E. orbicularis and M. leprosa on the one hand and additional hosts such as M. rivulata and the invasive turtle species Trachemys scripta (Thunberg) [ 19 ] on the other. Therefore, one may wonder whether the haemogregarine distribution is correlated to that of its definitive host, namely the Placobdella leech.…”

“…Due to the low host specificity of H. stepanowi against its intermediate host on the one hand [ 12 , 13 , 14 ] and the high diversity of Haemogregarina spp. in certain turtle species on the other [ 16 , 17 , 18 , 19 ], the host specificity of Haemogregarina could act against leeches—namely Placobdella as the definitive host—as claimed earlier by Attia El Hili et al [ 16 ].…”

“…Nine distinct COI haplotypes were revealed corresponding to three molecular lineages, each corresponding to a distinct Haemogregarina species, as previously advocated from an analysis of nuclear markers [ 15 ]. A similarly high Haemogregarina genetic diversity has since been evidenced from mitochondrial markers within the Savanna side-necked turtle Podocnemis vogli Müller [ 17 ], as well as from nuclear markers within the yellow-spotted Amazon river turtle Podocnemis unifilis Troschel [ 18 ] and E. orbicularis [ 19 ]. Due to the low host specificity of H. stepanowi against its intermediate host on the one hand [ 12 , 13 , 14 ] and the high diversity of Haemogregarina spp.…”

Eight-Year Study of Haemogregarina stepanowi Infection in Poached European Pond Turtles (Emys orbicularis) Held in Belgrade Zoo Quarantine

“…Molecular phylogenies have been based on nuclear, mitochondrial, and apicoplast DNA sequences [23]. The most recently published data indicate the presence of different lineages of Haemogregarinae in E. orbicularis obtained from seven different localities in Serbia and North Macedonia [24]. However, all animals monitored in our investigation were derived from the same natural habitat, and they were kept in the same pond in the quarantine section of Belgrade Zoo, which could explain the detection of the amplifications of 18S rRNA sequence in all 31 positive animals.…”