Citation: Ozubek, S.; Ulucesme, M.C.; Aktas, M. Discovery of a Novel Species Infecting Goats: Morphological and Molecular Characterization of Babesia aktasi n. sp. Pathogens 2023, 12, 113. https:// doi.org/10.3390/pathogens12010113 Academic Editors: Ana Domingos and Sandra Antunes Received: 2 December 2022 Revised: 1 January 2023 Accepted: 3 January 2023 Published: 10 January 2023 Copyright: © 2023 by the authors. Licensee MDPI, Basel, Switzerland. This article is an open access article distributed under the terms and conditions of the Creative Commons Attribution (CC BY) license (https:// creativecommons.org/licenses/by/ 4.0/). pathogens Article Discovery of a Novel Species Infecting Goats: Morphological and Molecular Characterization of Babesia aktasi n. sp. Sezayi Ozubek *, Mehmet Can Ulucesme and Munir Aktas Department of Parasitology, Faculty of Veterinary Medicine, University of Firat, Elazig 23200, Turkey * Correspondence: sozubek@firat.edu.tr Abstract: A novel Babesia sp. infecting goats was discovered based on the molecular findings obtained in the current study, which was conducted in the Mediterranean region of Türkiye. The goal of this study was to isolate this species of Babesia (Babesia sp.) infecting goats in vivo and to assess the genetic and morphological characterization of the parasite. To identify the animal naturally infected with Babesia sp. and isolate the parasite from this animal, field studies were conducted first, and genomic DNA were extracted from blood samples taken from goats (n = 50). The Theileria, Babesia, and Anaplasma species were identified using a nested PCR-based reverse line blotting (RLB) method. The study included one goat that was determined to be infected with Babesia sp. (single infection) in RLB for in vivo isolation. A blood smear was prepared to examine the parasite’s morphology, but it was found to be negative microscopically. Following that, a splenectomy operation (to suppress the immune system) was performed to make the parasites visible microscopically in this animal. Parasitemia began after splenectomy, and the maximum parasitemia was determined to be 1.9%. The goat displayed no significant symptoms other than fever, loss of appetite, and depression. During a period when parasitemia was high, blood from this goat was inoculated into another splenectomized goat (Theileria-Babesia-Anaplasma-Mycoplasma spp. free). On the third day of inoculation, 10% parasitemia with high fever was detected in the goat, and on the fourth day, the goat was humanely euthanized due to severe acute babesiosis symptoms. Except for mild subcutaneous jaundice, no lesions were discovered during the necropsy. According to the microscopic measurement results, ring, double pyriform, spectacle-frame-like, and line forms were observed, and it was observed to be between 1.0–2.5 μm (1.38 ± 0.17 to 0.7 ± 0.21-all forms). A phylogenetic analysis and sequence comparison using the 18S rRNA and cox1 genes revealed that this species is distinct from the small ruminant Babesia species (18S rRNA 92–94%, cox1 79–80%) and has the highest similarity to Babesia sp. deer, which has been reported in deer. Furthermore, it was determined to resemble B. venatorum, B. divergens, Babesia sp. FR1 and Babesia sp. MO1 species, all of which are zoonotic. Additional research is needed to clarify the clinical status of this parasite in goats and other hosts (mountain goat, sheep, calf). Keywords: Babesia; goat; morphology; phylogenetic analysis; sequence comparison 1. Introduction Shortly after the discovery of Babesia in bovine erythrocytes by Victor Babes in 1888, the same researcher reported that a parasite with similar characteristics also infected sheep [1]. Since then, it has been reported that Babesia ovis, B. motasi, B. crassa, B. taylori and B. foliate cause babesiosis in sheep and goats [2]. Babesiosis is a tick-borne disease caused by the genus Babesia, which is frequently observed in domestic and wild animals in tropical and subtropical regions [2–4]. The most pathogenic species is Babesia ovis, which is endemic to southern Europe, Africa, the Middle East, and Asia [5], and causes severe economic losses in sheep and goats [5]. Babesia crassa, which has low pathogenicity, was isolated in Iran, and B. motasi, which includes more than one species and subspecies, was isolated in China and Europe. B. motasi infections can cause mild clinical signs in sheep but can lead to severe Pathogens 2023, 12, 113. https://doi.org/10.3390/pathogens12010113 https://www.mdpi.com/journal/pathogens