Truly dorsostable runners: Vertebral mobility in rhinoceroses, tapirs, and horses.

The vertebral column is a hallmark of vertebrates; it is the structural basis of their body and the locomotor apparatus in particular. Locomotion of any vertebrate animal in its typical habitat is directly associated with functional adaptations of its vertebrae. This study is the first large-scale analysis of mobility throughout the presacral region of the vertebral column covering a majority of extant odd-toed ungulates from 6 genera and 15 species. In this study, we used a previously developed osteometry-based method to calculate available range of motion. We quantified all three directions of intervertebral mobility: sagittal bending (SB), lateral bending (LB), and axial rotation (AR). The cervical region in perissodactyls was found to be the most mobile region of the presacral vertebral column in LB and SB. Rhinoceroses and tapirs are characterized by the least mobile necks in SB among odd-toed and even-toed ungulates. Equidae are characterized by very mobile necks, especially in LB. The first intrathoracic joint (T1-T2) in Equidae and Tapiridae is characterized by significantly increased mobility in the sagittal plane compared to the typical thoracic joints and is only slightly less mobile than typical cervical joints. The thoracolumbar part of the vertebral column in odd-toed ungulates is very stiff. Perissodactyls are characterized by frequent fusions of vertebrae with each other with complete loss of mobility. The posterior half of the thoracic region in perissodactyls is characterized by especially stiff intervertebral joints in the SB direction. This is probably associated with hindgut fermentation in perissodactyls: the sagittal stiffness of the posterior thoracic region of the vertebral column is able to passively support the hindgut heavily loaded with roughage. Horses are known as a prime example of a dorsostable galloper among mammals. However, based on SB in the lumbosacral part of the backbone, equids appear to be the least dorsostable among extant perissodactyls; the cumulative SB in equids and tapirs is as low as in the largest representatives of artiodactyls, while in Rhinocerotidae it is even lower representing the minimum across all odd-toed and even-toed ungulates. Morphological features of small Paleogene ancestors of rhinoceroses and equids indicate that dorsostability is a derived feature of perissodactyls and evolved convergently in the three extant families.

Body size and digestive system shape resource selection by ungulates: A cross-taxa test of the forage maturation hypothesis.

The forage maturation hypothesis (FMH) states that energy intake for ungulates is maximised when forage biomass is at intermediate levels. Nevertheless, metabolic allometry and different digestive systems suggest that resource selection should vary across ungulate species. By combining GPS relocations with remotely sensed data on forage characteristics and surface water, we quantified the effect of body size and digestive system in determining movements of 30 populations of hindgut fermenters (equids) and ruminants across biomes. Selection for intermediate forage biomass was negatively related to body size, regardless of digestive system. Selection for proximity to surface water was stronger for equids relative to ruminants, regardless of body size. To be more generalisable, we suggest that the FMH explicitly incorporate contingencies in body size and digestive system, with small-bodied ruminants selecting more strongly for potential energy intake, and hindgut fermenters selecting more strongly for surface water.

Biogeography a key influence on distal forelimb variation in horses through the Cenozoic.

Locomotion in terrestrial tetrapods is reliant on interactions between distal limb bones (e.g. metapodials and phalanges). The metapodial-phalangeal joint in horse (Equidae) limbs is highly specialized, facilitating vital functions (shock absorption; elastic recoil). While joint shape has changed throughout horse evolution, potential drivers of these modifications have not been quantitatively assessed. Here, I examine the morphology of the forelimb metacarpophalangeal (MCP) joint of horses and their extinct kin (palaeotheres) using geometric morphometrics and disparity analyses, within a phylogenetic context. I also develop a novel alignment protocol that explores the magnitude of shape change through time, correlated against body mass and diet. MCP shape was poorly correlated with mass or diet proxies, although significant temporal correlations were detected at 0-1 Myr intervals. A clear division was recovered between New and Old World hipparionin MCP morphologies. Significant changes in MCP disparity and high rates of shape divergence were observed during the Great American Biotic Interchange, with the MCP joint becoming broad and robust in two separate monodactyl lineages, possibly exhibiting novel locomotor behaviour. This large-scale study of MCP joint shape demonstrates the apparent capacity for horses to rapidly change their distal limb morphology to overcome discrete locomotor challenges in new habitats.

Genetic diversity, evolution and selection in the major histocompatibility complex DRB and DQB loci in the family Equidae.

BACKGROUND: The mammalian Major Histocompatibility Complex (MHC) is a genetic region containing highly polymorphic genes with immunological functions. MHC class I and class II genes encode antigen-presenting molecules expressed on the cell surface. The MHC class II sub-region contains genes expressed in antigen presenting cells. The antigen binding site is encoded by the second exon of genes encoding antigen presenting molecules. The exon 2 sequences of these MHC genes have evolved under the selective pressure of pathogens. Interspecific differences can be observed in the class II sub-region. The family Equidae includes a variety of domesticated, and free-ranging species inhabiting a range of habitats exposed to different pathogens and represents a model for studying this important part of the immunogenome. While equine MHC class II DRA and DQA loci have received attention, the genetic diversity and effects of selection on DRB and DQB loci have been largely overlooked. This study aimed to provide the first in-depth analysis of the MHC class II DRB and DQB loci in the Equidae family. RESULTS: Three DRB and two DQB genes were identified in the genomes of all equids. The genes DRB2, DRB3 and DQB3 showed high sequence conservation, while polymorphisms were more frequent at DRB1 and DQB1 across all species analyzed. DQB2 was not found in the genome of the Asiatic asses Equus hemionus kulan and E. h. onager. The bioinformatic analysis of non-zero-coverage-bases of DRB and DQB genes in 14 equine individual genomes revealed differences among individual genes. Evidence for recombination was found for DRB1, DRB2, DQB1 and DQB2 genes. Trans-species allele sharing was identified in all genes except DRB1. Site-specific selection analysis predicted genes evolving under positive selection both at DRB and DQB loci. No selected amino acid sites were identified in DQB3. CONCLUSIONS: The organization of the MHC class II sub-region of equids is similar across all species of the family. Genomic sequences, along with phylogenetic trees suggesting effects of selection as well as trans-species polymorphism support the contention that pathogen-driven positive selection has shaped the MHC class II DRB/DQB sub-regions in the Equidae.

Equine Herpesvirus 1 Bridles T Lymphocytes To Reach Its Target Organs.

Equine herpesvirus 1 (EHV1) replicates in the respiratory epithelium and disseminates through the body via a cell-associated viremia in leukocytes, despite the presence of neutralizing antibodies. "Hijacked" leukocytes, previously identified as monocytic cells and T lymphocytes, transmit EHV1 to endothelial cells of the endometrium or central nervous system, causing reproductive (abortigenic variants) or neurological (neurological variants) disorders. In the present study, we questioned the potential route of EHV1 infection of T lymphocytes and how EHV1 misuses T lymphocytes as a vehicle to reach the endothelium of the target organs in the absence or presence of immune surveillance. Viral replication was evaluated in activated and quiescent primary T lymphocytes, and the results demonstrated increased infection of activated versus quiescent, CD4+ versus CD8+, and blood- versus lymph node-derived T cells. Moreover, primarily infected respiratory epithelial cells and circulating monocytic cells efficiently transferred virions to T lymphocytes in the presence of neutralizing antibodies. Albeit T-lymphocytes express all classes of viral proteins early in infection, the expression of viral glycoproteins on their cell surface was restricted. In addition, the release of viral progeny was hampered, resulting in the accumulation of viral nucleocapsids in the T cell nucleus. During contact of infected T lymphocytes with endothelial cells, a late viral protein(s) orchestrates T cell polarization and synapse formation, followed by anterograde dynein-mediated transport and transfer of viral progeny to the engaged cell. This represents a sophisticated but efficient immune evasion strategy to allow transfer of progeny virus from T lymphocytes to adjacent target cells. These results demonstrate that T lymphocytes are susceptible to EHV1 infection and that cell-cell contact transmits infectious virus to and from T lymphocytes.IMPORTANCE Equine herpesvirus 1 (EHV1) is an ancestral alphaherpesvirus that is related to herpes simplex virus 1 and causes respiratory, reproductive, and neurological disorders in Equidae. EHV1 is indisputably a master at exploiting leukocytes to reach its target organs, accordingly evading the host immunity. However, the role of T lymphocytes in cell-associated viremia remains poorly understood. Here we show that activated T lymphocytes efficiently become infected and support viral replication despite the presence of protective immunity. We demonstrate a restricted expression of viral proteins on the surfaces of infected T cells, which prevents immune recognition. In addition, we indicate a hampered release of progeny, which results in the accumulation of nucleocapsids in the T cell nucleus. Upon engagement with the target endothelium, late viral proteins orchestrate viral synapse formation and viral transfer to the contact cell. Our findings have significant implications for the understanding of EHV1 pathogenesis, which is essential for developing innovative therapies to prevent the devastating clinical symptoms of infection.

Fecal Carriage of Extended-Spectrum-ß-Lactamase/AmpC-Producing Escherichia coli in Horses.

A nationwide study on the occurrence of extended-spectrum ß-lactamase (ESBL)/AmpC in nonhospitalized horses in the Netherlands was performed. Molecular characterization was done, and questionnaires were analyzed to identify factors associated with carriage. In total, 796 horse owners were approached; 281 of these submitted a fecal sample from their horse(s), resulting in 362 samples. All samples were cultured qualitatively in Luria-Bertani (LB) broth and subsequently on MacConkey agar, both supplemented with 1 mg/liter cefotaxime (LB+ and MC+). Positive samples were subsequently cultured quantitatively on MC+. Initial extended-spectrum-ß-lactamase (ESBL)/AmpC screening was performed by PCR, followed by whole-genome sequencing on selected strains. Associations between ESBL/AmpC carriage and questionnaire items were analyzed using a univariate generalized estimating equation (GEE) regression analysis, followed by a multiple GEE model for relevant factors. In total, 39 of 362 samples (11%) were determined to be positive for ESBL/AmpC. blaCTX-M-1-carrying isolates were obtained from 77% of positive samples (n = 30). Other ESBL/AmpC genes observed included blaCTX-M-2, blaCTX-M-14, blaCTX-M-15, blaCTX-M-32, blaSHV-12, blaCMY-2, and blaACT-10 A high association between the presence of blaCTX-M-1 and IncHI1 plasmids was observed (46% of samples; n = 18). Based on core genome analysis (n = 48 isolates), six Escherichia coli clusters were identified, three of which represented 80% of the isolates. A negative association between ESBL/AmpC carriage and horses being in contact with other horses at a different site was observed. The presence of a dog on the premises and housing in a more densely human-populated region were positively associated.IMPORTANCE Extended-spectrum ß-lactamases (ESBLs) are widespread in human and animal populations and in the environment. Many different ESBL variants exist. The dissemination of ESBLs within and between populations and the environment is also largely influenced by genetic mobile elements (e.g., plasmids) that facilitate spread of these ESBLs. In order to identify potential attributable ESBL sources for, e.g., the human population, it is important to identify the different ESBL variants, the bacteria carrying them, and the potential risk factors for ESBL carriage from other potential sources. This nationwide study focuses on ESBL carriage in the open horse population and investigated the molecular characteristics, geographical distribution throughout the Netherlands, and potential risk factors for fecal ESBL carriage in horses. These data can be used for future attribution studies in order to reduce potential transmission of ESBL-producing bacteria between sources.

Purification of sarcoplasmic reticulum vesicles from horse gluteal muscle.

We have analyzed protein expression and enzyme activity of the sarcoplasmic reticulum Ca2+-transporting ATPase (SERCA) in horse gluteal muscle. Horses exhibit a high incidence of recurrent exertional rhabdomyolysis, with myosolic Ca2+ proposed, but yet to be established, as the underlying cause. To better assess Ca2+ regulatory mechanisms, we developed an improved protocol for isolating sarcoplasmic reticulum (SR) vesicles from horse skeletal muscle, based on mechanical homogenization and optimized parameters for differential centrifugation. Immunoblotting identified the peak subcellular fraction containing the SERCA1 protein (fast-twitch isoform). Gel analysis using the Stains-all dye demonstrated that calsequestrin (CASQ) and phospholipids are highly enriched in the SERCA-containing subcellular fraction isolated from horse gluteus. Immunoblotting also demonstrated that these horse SR vesicles show low content of glycogen phosphorylase (GP), which is likely an abundant contaminating protein of traditional horse SR preps. The maximal Ca2+-activated ATPase activity (Vmax) of SERCA in horse SR vesicles isolated using this protocol is 5‒25-fold greater than previously-reported SERCA activity in SR preps from horse skeletal muscle. We propose that this new protocol for isolating SR vesicles will be useful for determining enzymatic parameters of horse SERCA with high fidelity, plus assessing regulatory effect of SERCA peptide subunit(s) expressed in horse muscle.

Perissodactyla (Rhinocerotidae and Equidae) from Kanapoi.

The Kanapoi collection of Rhinocerotidae, first studied by Hooijer and Patterson (1972), now consists of 25 specimens and substantial reinterpretation of their affinities is made here. Kanapoi post-dates the extinction of Brachypotherium and the whole collection belongs to the Dicerotini. It is important because it includes the type-specimen of Diceros praecox, a species that remains poorly known, but looks slightly larger and more primitive than the modern 'black' rhino, Diceros bicornis. A second species is probably ancestral to the modern 'white' rhino, Ceratotherium simum; it looks identical to the Pleistocene North African Ceratotherium mauritanicum, of which Ceratotherium efficax is probably a synonym. The evolution of the Dicerotini in Africa can be regarded as an increasing divergence in diet and related morphofunctional adaptations in the two lineages. The co-occurrence at Kanapoi of both Diceros and Ceratotherium, with distinct dietary preferences, suggests some habitat heterogeneity, although the low sample size prevents robust paleoecological conclusions. The Equidae are also rare and consist mostly of isolated teeth. I take the most parsimonious option of tentatively including all of them in a single species, whose identification is left open. Dental features of eastern African Pliocene to Pleistocene hipparions may reflect increasing adaptation to grazing.

Bloodmeal analysis in Culicoides midges collected near horses, donkeys and zebras in the Eastern Cape, South Africa.

An upsurge in African horse sickness (AHS) in the Eastern Cape, South Africa, from 2006 led to an epidemiological reassessment of the disease there. Light trapping surveys carried out near horses, donkeys and zebras in 2014-2016 collected 39 species of Culicoides midge (Diptera: Ceratopogonidae) that are potential vectors of AHS. To establish if these midges fed on equids, DNA sequences were obtained from the gut contents of 52 female midges (35 freshly blood-fed, 13 gravid and four parous), representing 11 species collected across 11 sites. Culicoides leucostictus fed on all three equids. Culicoides bolitinos, Culicoides imicola and Culicoides magnus fed on both horses and donkeys. Culicoides onderstepoortensis fed on donkeys, and Culicoides similis and Culicoides pycnostictus fed on zebras. Bloodmeals from cows, pigs, warthogs, impalas and a domestic dog were also identified in various species, but none of the midges tested had fed on birds. These results contribute to knowledge of the vectorial capacity of several species of Culicoides with regard to AHS in the Eastern Cape and point to potential reservoir hosts, of which donkeys, zebras and domestic dogs have previously been found to harbour AHS. Blood-fed midges were also obtained throughout winter, indicating the potential for endemic AHS in the province.

African Horse Sickness Virus: History, Transmission, and Current Status.

African horse sickness virus (AHSV) is a lethal arbovirus of equids that is transmitted between hosts primarily by biting midges of the genus Culicoides (Diptera: Ceratopogonidae). AHSV affects draft, thoroughbred, and companion horses and donkeys in Africa, Asia, and Europe. In this review, we examine the impact of AHSV critically and discuss entomological studies that have been conducted to improve understanding of its epidemiology and control. The transmission of AHSV remains a major research focus and we critically review studies that have implicated both Culicoides and other blood-feeding arthropods in this process. We explore AHSV both as an epidemic pathogen and within its endemic range as a barrier to development, an area of interest that has been underrepresented in studies of the virus to date. By discussing AHSV transmission in the African republics of South Africa and Senegal, we provide a more balanced view of the virus as a threat to equids in a diverse range of settings, thus leading to a discussion of key areas in which our knowledge of transmission could be improved. The use of entomological data to detect, predict and control AHSV is also examined, including reference to existing studies carried out during unprecedented outbreaks of bluetongue virus in Europe, an arbovirus of wild and domestic ruminants also transmitted by Culicoides.

Effect of dietary plant extract on meat quality and sensory parameters of meat from Equidae.

BACKGROUND: Plant extracts as Lippia spp. have been proven antioxidant properties. Recent studies have been shown that dietary supplementation with plant extracts is able to enhance meat quality parameters. Studies regarding meat quality in Equidae are limited. RESULTS: The effect of dietary plant extract (PE), containing verbascoside, on meat quality, oxidative stability and sensory parameters of Longissimus Lumborum (LL) muscle in Equidae was studied. Dietary treatment did not affect (P > 0.05) pH, colour indices and chemical parameters of muscle in both donkey and horse. Dietary PE improved (P < 0.01) oxidative stability in donkey muscle during refrigerated storage. Sensory characteristics of LL muscle were positively affected (P < 0.05) by dietary PE in both donkey and horse. In particular, colour, taste and texture were enhanced in LL muscle from animals fed PE. Oxidative stability was lower (P < 0.05) in LL muscle of horse than that of donkey. CONCLUSION: Dietary plant extract, containing verbascoside, can be considered as a natural source of antioxidants, and is also able to improve oxidative stability of donkey meat and to affect the sensory attributes of Equidae meat. © 2017 Society of Chemical Industry.

DESCRIPTION OF GASTRIC ULCERS AND OF THEIR SUSPECTED, ASSOCIATED RISK FACTORS IN DECEASED WILD EQUIDS AT THE RÉSERVE AFRICAINE DE SIGEAN, FRANCE (2010-2016).

Gastric ulcers are common in domestic horses and foals, affecting at least 90% of unmedicated racehorses in active training. Despite these high prevalences in domestic horses, literature about this condition in wild equids is almost nonexistent. The presence of gastric ulcers was evaluated at necropsy in six species of wild equids that died at the Réserve Africane de Sigean, a safari park in the south of France from 2010 to 2016. Among the 55 individuals that died during that period, a description of the gastric mucosa was available in 82% (45/55) of cases. Considering the cases for which a description of the gastric mucosa was available, the prevalence of gastric ulcers was 64% (29/45). The highest prevalences were noted in Grant's zebra (Equus quagga boehmi) and Hartmann's mountain zebra (Equus zebra hartmannae) at 83% and 100%, respectively. In contrast to what is reported in domestic foals, gastric ulcerations were only diagnosed in one foal (out of 11 foals necropsied). The higher prevalence was noted in young individuals (3-36 mo old) at 93% (14/15); the lesions observed consisted mainly of single to multiple, superficial lesions, of which, only the mucosa was missing; these superficial lesions are often considered not clinically significant. The prevalence was lower for adults (74%; 14/19), but lesions were deeper or with a hyperemic or inflammatory appearance. All the lesions observed were located in the gastric, nonglandular, stratified squamous mucosa, along the margo plicatus. No statistical correlation could be found between the development of gastric ulcers and an ongoing, chronic pathologic process or a digestive tract pathology. The detection of gastric ulcers was, therefore, significantly greater in wild equids isolated in smaller enclosures. Nevertheless, additional larger-scale research is needed to point out predisposing factors in equids under human care.

Positive selection in the SLC11A1 gene in the family Equidae.

Immunity-related genes are a suitable model for studying effects of selection at the genomic level. Some of them are highly conserved due to functional constraints and purifying selection, while others are variable and change quickly to cope with the variation of pathogens. The SLC11A1 gene encodes a transporter protein mediating antimicrobial activity of macrophages. Little is known about the patterns of selection shaping this gene during evolution. Although it is a typical evolutionarily conserved gene, functionally important polymorphisms associated with various diseases were identified in humans and other species. We analyzed the genomic organization, genetic variation, and evolution of the SLC11A1 gene in the family Equidae to identify patterns of selection within this important gene. Nucleotide SLC11A1 sequences were shown to be highly conserved in ten equid species, with more than 97 % sequence identity across the family. Single nucleotide polymorphisms (SNPs) were found in the coding and noncoding regions of the gene. Seven codon sites were identified to be under strong purifying selection. Codons located in three regions, including the glycosylated extracellular loop, were shown to be under diversifying selection. A 3-bp indel resulting in a deletion of the amino acid 321 in the predicted protein was observed in all horses, while it has been maintained in all other equid species. This codon comprised in an N-glycosylation site was found to be under positive selection. Interspecific variation in the presence of predicted N-glycosylation sites was observed.

CRYOPRESERVATION OF ONAGER (EQUUS HEMIONUS ONAGER) EPIDIDYMAL SPERMATOZOA.

Genetic diversity is a primary component of adaptive evolution, and its loss or reduction can decrease the long-term survival probability of populations. Utilization of cryopreserved semen may be considered a perfect tool to improve genetic diversity, reduce inbreeding, and avoid animal translocation for breeding. The present study aimed at finding a reliable epididymal sperm freezing protocol for the critically endangered onager (Equus hemionus onager). Six testicles from three animals were processed postmortem. The effects of two transportation temperatures (22°C and 4°C; testicles submerged in saline), two cryopreservation techniques (conventional liquid nitrogen vapor freezing in straws and directional freezing in 8-ml HollowTubes(TM)), and two postthaw incubation temperatures (22°C and 37°C; evaluated after 0.5, 1, 2, and 3 hr) were tested in a 2×2×2 experimental design. Sperm samples were evaluated for motility, viability, acrosome integrity, and sperm morphology. The resulting optimal freezing protocol includes transportation of testicles at 4°C, cryopreservation by directional freezing, and, if needed, postthaw incubation at 22°C. With this combination of transportation temperature and cryopreservation technique, the authors obtained the following postthaw values normalized to prefreezing values: 60.3±8.8% motility, 60.7±13.3% viability, 75.3±9.5% acrosome integrity, and 94.7±2.9% normal morphology (excluding defects due to the epididymal origin of the sperm). After incubation at 22°C, motility values for the above combination were 40±5.7%, 30.3±5.2%, 28.3±4.4%, and 16.7±4.4% for 0.5, 1, 2, and 3 hr, respectively. In conclusion, with this protocol, good quality semen can be stored for future use in artificial inseminations when and where needed.

Body mass evolution and diversification within horses (family Equidae).

Horses (family Equidae) are a classic example of adaptive radiation, exhibiting a nearly 60-fold increase in maximum body mass and a peak taxonomic diversity of nearly 100 species across four continents. Such patterns are commonly attributed to niche competition, in which increased taxonomic diversity drives increased size disparity. However, neutral processes, such as macroevolutionary 'diffusion', can produce similar increases in disparity without increased diversity. Using a comprehensive database of Equidae species size estimates and a common mathematical framework, we measure the contributions of diversity-driven and diffusion-driven mechanisms for increased disparity during the Equidae radiation. We find that more than 90% of changes in size disparity are attributable to diffusion alone. These results clarify the role of species competition in body size evolution, indicate that morphological disparity and species diversity may be only weakly coupled in general, and demonstrate that large species may evolve from neutral macroevolutionary diffusion processes alone.

Ere, a Family of Short Interspersed Elements in the Genomes of Odd-Toed Ungulates (Perissodactyla).

Short Interspersed Elements (SINEs) are eukaryotic retrotransposons transcribed by RNA polymerase III (pol III). Many mammalian SINEs (T+ SINEs) contain a polyadenylation signal (AATAAA), a pol III transcription terminator, and an A-rich tail in their 3'-end. The RNAs of such SINEs have the capacity for AAUAAA-dependent polyadenylation, which is unique to pol III-generated transcripts. The structure, evolution, and polyadenylation of the Ere SINE of ungulates (horses, rhinos, and tapirs) were investigated in this study. A bioinformatics analysis revealed the presence of up to ~4 × 105 Ere copies in representatives of all three families. These copies can be classified into two large subfamilies, EreA and EreB, the former distinguished by an additional 60 bp sequence. The 3'-end of numerous EreA and all EreB copies exhibit a 50 bp sequence designated as a terminal domain (TD). The Ere family can be further subdivided into subfamilies EreA_0TD, EreA_1TD, EreB_1TD, and EreB_2TD, depending on the presence and number of terminal domains (TDs). Only EreA_0TD copies can be assigned to T+ SINEs as they contain the AATAAA signal and the TCTTT transcription terminator. The analysis of young Ere copies identified by comparison with related perissodactyl genomes revealed that EreA_0TD and, to a much lesser extent, EreB_2TD have retained retrotranspositional activity in the recent evolution of equids and rhinoceroses. The targeted mutagenesis and transfection of HeLa cells were used to identify sequences in equine EreA_0TD that are critical for the polyadenylation of its pol III transcripts. In addition to AATAAA and the transcription terminator, two sites in the 3' half of EreA, termed the ß and τ signals, were found to be essential for this process. The evolution of Ere, with a particular focus on the emergence of T+ SINEs, as well as the polyadenylation signals are discussed in comparison with other T+ SINEs.

First report of infection with metacestode stages of Echinococcus multilocularis in a kulan (Equus hemionus kulan) from Slovakia.

While the principle definitive host of the zoonotic cestode Echinococcus multilocularis in Europe is the red fox, several rodent species act as main intermediate hosts. Among others, e.g., humans, dogs, and pigs, also horses have been described to act as aberrant hosts in highly endemic regions. Here, a case of an E. multilocularis infection in a kulan (Equus hemionus kulan) is described. The five years old kulan from a zoo in Slovakia was transported to an animal park in Germany. The animal had to be euthanized within a few weeks after the import due to its poor general state of health. The pathological examination revealed a nodular mass in the liver as an incidental finding. By histological examination of the mass, a pyogranulomatous and necrotizing inflammation and intralesional fragments of amorphous eosinophil layers were detected. The suspected diagnosis of E. multilocularis infection was confirmed by PCR addressing parts of the genes 12S rRNA and the NADH dehydrogenase subunit 2, showing very high identities with isolates from France, Slovakia and the USA.

Relevant Brachycera (Excluding Oestroidea) for Horses in Veterinary Medicine: A Systematic Review.

In equine stables and their surroundings, a large number of insects are present that can be a nuisance to their equine hosts. Previous studies about dipterans transmitting infectious agents to Equidae have largely focused on Nematocera. For the preparation of this systematic review, the existing literature (until February 2022) was systematically screened for various infectious agents transmitted to Equidae via insects of the suborder Brachycera, including Tabanidae, Muscidae, Glossinidae and Hippoboscidae, acting as pests or potential vectors. The PRISMA statement 2020 (Preferred Reporting Items for Systematic Reviews and Meta-Analyses) guidelines for systematic reviews were followed. The two concepts, Brachycera and Equidae, were combined for the search that was carried out in three languages (English, German and French) using four different search engines. In total, 38 articles investigating Brachycera as vectors for viral, bacterial and parasitic infections or as pests of equids were identified. Only 7 of the 14 investigated pathogens in the 38 reports extracted from the literature were shown to be transmitted by Brachycera. This review clearly shows that further studies are needed to investigate the role of Brachycera as vectors for pathogens relevant to equine health.

Strong Evidence of the Role of Donkeys in the Epidemiology of Leptospira spp. in Semiarid Conditions.

Donkeys (Equus asinus) are historically known for their close relationship to humanity, which raises the need to study zoonotic diseases that affect them. In this perspective, leptospirosis stands out as a disease with an economic and public health impact, and its occurrence is facilitated in times of higher rainfall indexes, especially in large urban centers. In view of the scarcity of information about leptospirosis in donkeys, the objective of this study was to detect the presence of Leptospira spp. and anti-leptospiral antibodies in donkeys rescued by a zoonosis center located in the Caatiga biome, Brazilian semiarid region. Overall, 30 donkeys of both sexes, aged between 4 months and 15 years, were used, from which 64 serum samples were collected and submitted to the microscopic agglutination test (MAT). In addition, 64 samples of urine, vaginal and preputial fluid, in duplicates, were subjected to the polymerase chain reaction (PCR) and microbiological. Sixteen (53.3%) animals tested positive in at least one diagnostic test, 12 (40%) of which were positive at MAT and seven (23.3%) in the molecular and bacteriological detection (urine, vaginal, and preputial fluid samples). This is the first report identifying donkeys infected with Leptospira spp. by molecular and bacteriological diagnosis in Brazil, and the first in the world to detect this agent in their genital fluids. The study also shows that donkeys are commonly exposed to leptospires in the Caatinga biome, and this constitutes a One Health-based concern, demonstrating the importance of broad studies where large numbers of humans and animals coexist when investigating zoonotic infections and when planning and implementing control measures for donkeys-associated leptospirosis.

Equine Rolling Behavior: Thermoregulation Mechanism After Exercise and Substrate Preference.

Rolling is a natural behavior of equids that is beneficial to their well-being. However, more intensive domestic environments offer fewer opportunities for animals to roll, neglecting its importance. We believe that the inclusion of recreational areas for animals, containing substrates and conditions that encourage the occurrence of natural behaviors, is essential to promote their well-being. Therefore, we evaluated the preference of horses (Equus caballus; n = 8) and mules (E. asinus x E. caballus; n = 8) for different rolling substrates (sand, grass, manure), and the influence of this behavior on thermoregulation after exercise. Horses rolled preferentially on grass, whereas mules preferred sand, both of which were cooler substrates when compared to manure. In addition, the temperature of the animals significantly reduced after rolling. Thus, horses and mules must roll for thermoregulation after exercise, choosing cooler and more efficient substrates for heat dissipation. The differences in preferences between mules and horses showed particularities between the equine species and its hybrid, which is important for management that is focused more on the individual animal.