Unoculubranchiobdella sp. (Hirudinea: Ozobranchidae) as a vector for Haemogregarina spp. in freshwater turtles from Brazil.

Species of Haemogregarina are blood parasites known to parasitise vertebrate hosts, including fishes (Haemogregarina sensu lato) and freshwater turtles (Haemogregarina sensu stricto). Their vectors, include gnathiid isopods and leeches, respectively. In turtles, Haemogregarina balli has the best-characterized life cycle in the genus. However, no studies in Brazil have suggested a possible vector for any species of Haemogregarina from freshwater turtles. Therefore, in the present study, we provide insights into a leech vector based on specimens found feeding on two species of freshwater turtles, Podocnemis unifilis and Podocnemis expansa, using morphological and molecular data. In 2017 and 2019, freshwater turtles were collected in Goiás State, Brazil. Hosts were inspected for ectoparasites and leeches were collected from two specimens of P. expansa and nine specimens of P. unifilis. Leeches were subsequently identified as members of the genus Unoculubranchiobdella. Leech histological slides revealed haemogregarine-like structures, similar to post-sporogonic merogony, found near the gills and within the posterior sucker. Molecular analysis of the haemeogregarines resulted in the identification of three species of Haemogregarina: Haemogregarina embaubali, Haemogregarina goianensis, and Haemogregarina brasiliana. Therefore, our findings, based on morphology and DNA data suggest leeches of the genus Unoculubranchiondella as vectors for at least three species of Haemogregarina from Brazilian turtles.

Closing the knowledge gap: Helminth parasites of freshwater turtles from the Chaco-Pampa Plain, Southern South America.

Six species of freshwater turtles dominate the Chaco-Pampa Plain in southern South America and their parasites have been relatively understudied, with most records concentrated in Brazil. Particularly in Argentina, there are only scattered records of parasites for most of the turtles that inhabit the region, leaving a large knowledge gap. The purpose of the present contribution is to increase the knowledge of the internal parasites of six species of freshwater turtles from Argentina, after 15 years of fieldwork, by providing new hosts and additional geographic records for many host-parasite relationships. Some molecular sequences of the studied parasites were provided as a tool for better species identification. We processed 433 stomach and fecal samples from live individuals and visceral and soft tissue samples from 54 dissected turtles collected from a wide range and different ecoregions. We found 6230 helminths belonging to 18 taxa (one cestode, 11 digeneans and six nematodes). Fourteen new parasite-host associations are reported here, and for the first time parasites are recorded for Phrynops williamsi. This work contributes significantly to the knowledge of the parasitofauna in freshwater turtles in Argentina, providing a detailed list of parasites present in each turtle species and reporting molecular characters for future studies.

Orientatractis bowseri n. sp. (Nematoda: Cosmocercoidea: Atractidae) parasite of Peltocephalus dumerilianus (Schweigger) (Testudines: Podocnemididae) from the Brazilian Amazon.

Peltocephalus dumerilianus (Schweigger) is a diurnal freshwater turtle widely distributed in the Orinoco and Amazon River basins in Brazil, Colombia, Peru, Ecuador, Venezuela, and French Guiana. During a helminthological survey of freshwater turtles from the Brazilian Amazon, numerous nematodes with characteristics of the genus Orientatractis Petter, 1966 were recovered from the stomach of Peltocephalus dumerilianus. These nematodes were different from all congeneric species, and we describe herein. Orientatractis bowseri n. sp. differs from all known species of the genus by having dorsal cuticular projections on the posterior region of the females, distribution of caudal papillae and by having papilla-like ornamentations on the cuticle distributed from the mid-esophagus to the mid portion of the body. This study represents the fourth specie of the genus Orientatractis reported in Brazil, the seventh species of Orientatractis in the Neotropical region, and the third found infecting freshwater turtles in Brazil.

Pathological changes by spirorchiid eggs in hawksbill sea turtle (Eretmochelys imbricata) stranded off brazilian coast.

Eretmochelys imbricata (hawksbill turtle) is classified as a critically endangered species at world level; in Brazil, it is listed among the threatened species. Spirorchiids are parasites of the circulatory system of chelonians which may cause serious lesions in the various tissues of the host due to deposition of eggs in the bloodstream. In this context, the aim of the present study was to describe the pathology caused by spirorchiid eggs in E. imbricata from the Brazilian over a five year period. A total of 29 animals were analyzed, of which nine (31%) presented lesions associated to spirorchiids eggs. The lesions were: granulomatous enteritis in six (66.66%), granulomatous splenitis in five (55.55%), granulomatous hepatitis in three (33.33%), granulomatous pneumonia in three (33.33%), granulomatous pancreatitis in two (22.22%), and granulomatous adrenalitis in one specimen (11.11%). Concluded the main lesion in Eretmochelys imbricata was giant-cell granulomatous inflammatory reaction to the parasite's eggs. Animals exhibiting poor physical health were more susceptible to presenting such lesions.

Blood parasite diversity (Apicomplexa: Haemogregarinidae) within the western populations of the European pond turtle Emys orbicularis.

Molecular tools have revolutionized assessments of blood parasites in freshwater turtles. In the Iberian Peninsula and North Africa, two native species of terrapins occur, Emys orbicularis (Linnaeus) and Mauremys leprosa (Schweigger). Both have been identified as hosts for the blood parasite Haemogregarina stepanowi Danilewsky, 1885, which has also been found in related species. However, recent assessments of M. leprosa have identified several distinct genetic lineages of these parasites in this host, while only three haemogregarine lineages were identified in E. orbicularis in Tunisia. Here, we screened 215 individuals of E. orbicularis from the Iberian Peninsula, Menorca Island and Morocco for haemogregarine parasites using partial 18S rRNA gene sequences to estimate relationships. Three unrelated lineages of parasites were detected, one presumed H. stepanowi and two lineages previously known from M. leprosa. A considerable undescribed diversity of parasites exists within these vertebrate host species, while mixed infection and host-sharing is also widespread. Considering that E. orbicularis is near threatened in this region, it is of great importance to identify the parasites infecting it, and to further assess the potential deleterious impact of these diverse parasites on their hosts.

Vogtnema asymmetrica n. gen., n. sp. (Nematoda: Cosmocercoidea: Atractidae) parasite of Rhinoclemmys punctularia (Daudin) (Testudines: Geoemydidae) from eastern Amazon, Brazil.

Nematodes collected from the stomach and large intestine of Rhinoclemmys punctularia (Daudin) from the eastern Amazon, Brazil, are assigned to a new genus, Vogtnema n. gen., allocated in the family Atractidae (Cosmocerdoidea). Vogtnema n. gen. differs from all other genera of Atractidae based on the combination of the following morphological characters: four lips V-shaped (two subdorsal and two subventral) without sclerotized structures associated, absence of sclerotized formations and/or pieces surrounding the oral opening, oral aperture surrounded by flange internal membranous structures; in males the larger spicule is alate. The type species Vogtnema asymmetrica n. gen., n. sp. has the five precloacal pairs of papillae situated on the right side of the body that are more prominent than those on the left side. We also provide an identification key to the genera of the family Atractidae.

Molecular and morphological description of the first Hepatozoon (Apicomplexa: Hepatozoidae) species infecting a neotropical turtle, with an approach to its phylogenetic relationships.

Haemogregarines (Adeleorina) have a high prevalence in turtles. Nevertheless, there is only one Hepatozoon species described that infects Testudines so far; it is Hepatozoon fitzsimonsi which infects the African tortoise Kinixys belliana. Colombia harbours a great diversity of chelonians; however, most of them are threatened. It is important to identify and characterize chelonian haemoparasite infections to improve the clinical assessments, treatments and the conservation and reintroduction programs of these animals. To evaluate such infections for the Colombian wood turtle Rhinoclemmys melanosterna, we analysed blood from 70 individuals. By using the morphological characteristics of blood stages as well as molecular information (18S rRNA sequences), here we report a new Hepatozoon species that represents the first report of a hepatozoid species infecting a semi-aquatic continental turtle in the world. Although the isolated lineage clusters within the phylogenetic clades that have morphological species of parasites already determined, their low nodal support makes their position within each group inconclusive. It is important to identify new molecular markers to improve parasite species identification. In-depth research on blood parasites infecting turtles is essential for increasing knowledge that could assess this potential unknown threat, to inform the conservation of turtles and for increasing the state of knowledge on parasites.

Health assessment of wild speckled dwarf tortoises, CHERSOBIUS SIGNATUS.

BACKGROUND: In free-ranging reptile populations, bacterial, fungal, viral and parasitic pathogens may affect hosts through impairment in movements, thermoregulation, reproduction, survival, and population dynamics. The speckled dwarf tortoise (Chersobius [Homopus] signatus) is a threatened species that is mostly restricted to the Succulent Karoo biome in South Africa, and little information on pathogens of this species is available yet. We derived baseline parameters for five males and five females that were captured to genetically enhance a conservation breeding program in Europe. Upon collection of the tortoises, ticks were removed and identified. Immediately upon arrival in Europe, ocular, nasal, oral and cloacal swabs were taken for viral, bacteriological and mycological examinations. Fecal samples were collected before and 1 month after fenbendazole treatment, and analyzed for parasites. A panel of PCR, aiming to detect herpesviruses, adenoviruses and iridoviruses, was carried out. RESULTS: Samples were negative for viruses, while bacteriological examination yielded detectable growth in 82.5% of the swabs with a mean load of 16 × 107 ± 61 × 108 colony forming units (CFU) per swab, representing 34 bacterial species. Cloacal and oral swabs yielded higher detectable growth loads than nasal and ocular swabs, but no differences between sexes were observed. Fungi and yeasts (mean load 5 × 103 ± 13 × 103 CFU/swab) were detected in 25% of the swabs. All pre-treatment fecal samples were positive for oxyurid eggs, ranging from 200 to 2400 eggs per gram of feces, whereas after the treatment a significantly reduced egg count (90-100% reduction) was found in seven out of 10 individuals. One remaining individual showed 29% reduction, and two others had increased egg counts. In five tortoises, Nycthocterus spp. and coccidian oocysts were also identified. Soft ticks were identified as Ornithodoros savignyi. CONCLUSIONS: Our baseline data from clinically healthy individuals will help future studies to interpret prevalences of microorganisms in speckled dwarf tortoise populations. The study population did not appear immediately threatened by current parasite presence.

Desmogonius baldassinae n. sp. (Digenea: Pronocephalidae) collected in a green sea turtle-Chelonia mydas-from Brazil.

The genus Desmogonius (Digenea: Pronocephalidae) is known only from sea turtles and currently contains two species: D. desmogonius Stephens, 1911 and D. loossi Chattopadhyaya, 1972. The present study describes a third species, Desmogonius baldassinae n. sp. found in a juvenile green sea turtle (Chelonia mydas) from Brazil. The new species is larger than its congeners in all dimensions, with vitellinic fields ending posterior to the ovary. In addition, we present a key for species identification and an emended diagnosis of the genus Desmogonius.

Cucullanus carettae Baylis, 1923, in a loggerhead sea turtle (Caretta caretta) from the Adriatic sea: first detection and molecular characterization.

Cucullanus carettae Baylis, 1923 (Nematoda: Cucullanidae) is found worldwide in loggerhead turtles (Caretta caretta). Regarding the Mediterranean, C. carettae has been identified in the Tyrrhenian and the Ionian Sea and a unique description of a Cucullanus sp. specimen in loggerheads from the Adriatic Sea has been reported in the literature so far. In the framework of a bio-monitoring project of the Abruzzo and Molise coasts, a parasitological survey was performed on stranded and by-caught sea turtles, at the Istituto Zooprofilattico of Abruzzo and Molise "G. Caporale." During necropsy, the gastrointestinal system of 72 stranded loggerhead turtles was analyzed for the presence of endoparasites and fecal samples were collected for coprological examination. Adult C. carettae (n = 123) was found in the upper intestine of one loggerhead turtle, associated with chronic lymphoplasmocytic enteritis. Additionally, five stool samples (6.9%) were positive for Cucullanus sp. eggs. Molecular characterization of adult nematodes was carried out to study phylogenetic relationships among the Cucullanus species. To our knowledge, this is the first morphological and molecular identification of C. carettae in loggerhead turtles from the Adriatic Sea. Additional studies on the distribution of this parasite in the Mediterranean are encouraged.

Molecular evidence of new freshwater turtle blood flukes (Digenea: Spirorchiidae) in the intermediate snail host Biomphalaria occidentalis Paraense, 1981 in an urban aquatic ecosystem in Brazil.

In this study, two potentially new species of turtle blood flukes (TBFs) (Digenea: Spirorchiidae) have been recorded from South Brazil. The spirorchiid parasites infect the vascular system of turtles, thereby compromising their health. The life cycle of these parasites is not well studied. The larval stage of cercaria is found in intermediate gastropod hosts, with some species presenting similar morphological characteristics, which can result in misinterpretations when using only morphological taxonomy for species identification. In this study, we recorded a single morphotype belonging to the family Spirorchiidae in Biomphalaria occidentalis in an urban aquatic ecosystem in Brazil. However, molecular data (28S rDNA and cytochrome oxidase subunit I) confirmed the presence of two species of Spirorchiidae in the sampled environment; both phylogenetically close to genera previously studied in freshwater turtles from the Peruvian Amazon. In this study, species characterization was possible because of molecular tools. We recommend using more than one molecular marker in future studies focusing on TBFs, which need attention about their evolutionary history and ecology to understand their distribution in South America.

Characterization and identification of haemogregarine hemoparasites (Apicomplexa: Adeleina: Hepatozoidae) in natural populations of Mauremys leprosa leprosa and M. leprosa saharica from Morocco.

Distribution, prevalence and parasitaemia of apicomplexan parasites of the genus Haemogregarina were studied in 858 freshwater turtles (735 Mauremys leprosa leprosa and 123 Mauremys leprosa saharica) throughout 30 localities from Morocco. Blood smears were collected from the turtles and partial 18S rRNA sequence data used to infer genetic diversity and phylogenetic relationships. Of the 858 individuals analyzed by microscopy 22.7% were infected, from 16 M. leprosa leprosa localities. Individuals of M. leprosa saharica sampled south of the High Atlas Mountains were not infected, probably due to the absence or rarity of the leech vectors in these localities. Within M. leprosa leprosa, we did not identify any patterns between geography and prevalence, which varied between 10% (Oued Nfiss) to 100% (Oued Tassaout). Five distinct genetic lineages were identified, indicating the likely presence of multiple species of haemogregarines, one of which probably corresponds to Haemogregarina stepanowi. Mixed infections were also detected. Additional studies are needed to better understand the ecology and epidemiology of this parasite in turtles, as well as the host-parasite relationship with their definitive hosts, leeches.

Placobdelloides tridens sp. n., a new species of glossiphoniid leech (Hirudinea: Rhynchobdellida) found feeding on captive Orlitia borneensis in Thailand, and an update to the host distribution of P. siamensis.

A new glossiphoniid leech species, Placobdelloides tridens sp. n., is discovered on the Malayan Giant Turtle (Orlitia borneensis) at the Nakhon Ratchasima Zoo in Thailand. The morphological study of this new species revealed that it is distinguished from P. siamensis, a turtle leech species that can be found commonly in Thailand. Placobdelloides tridens presented the following diagnostic morphological characteristics: a pear-shaped and triannulate body, well-developed rod-like papillae on the dorsal surface, smooth posterior and anterior suckers with nominal pits inside, a single pair of dark contiguous eyes, light yellow-brown to greenish dorsal color, absence of median line, male and female gonopore separated by a single annulus and a unique trident shape at the tip of the crop ceca. The phylogenetic relationships of P. tridens sp. n., was clarified, and shown to be a sister clade to the P. siamensis and P. sirikanchanae clade. Furthermore, this is a new host record for P. siamensis, which was found on O. borneensis, Batagur affinis and B. borneoensis in the Khao Kheow Open Zoo, Chonburi, Thailand.

A pilot study on molecular diagnosis of Hapalotrema mistroides (Digenea: Spirorchiidae) infection in blood samples of live loggerhead turtles Caretta caretta.

BACKGROUND: Parasites of the family Spirorchiidae cause disease and mortality in marine and freshwater turtles; two species, Hapalotrema mistroides and Neospirorchis sp., are reported in the resident population of loggerhead turtles of the Mediterranean Sea, with the first being the most widespread. In vivo diagnosis of spirorchidiasis can represent a challenge in guaranteeing prompt control and treatment of the disease and is currently limited to copromicroscopy. The aim of this study was the development of a real time PCR assay with TaqMan probe for the detection of H. mistroides infection in the blood of live loggerhead turtles, Caretta caretta, hospitalized in rehabilitation centres. Its potential use for in vivo diagnosis is explored. RESULTS: The developed real time PCR successfully detected H. mistroides DNA from both positive controls and experimental blood samples of live loggerhead sea turtles, showing good specificity, sensitivity and good reaction efficiency. Two out of three turtles which had demonstrated positivity at copromicroscopy also tested positive to this blood assay; DNA of H. mistroides was detected within the blood of one sea turtle, which tested negative for copromicroscopy. CONCLUSIONS: This study describes a specific and rapid molecular assay to detect H. mistroides infection from live sea turtles and highlights for the first time the presence of DNA of this species in turtle blood samples. Since this assay is able to detect low amounts of the parasitic free DNA in blood samples, its application could be helpful for in vivo diagnosis of H. mistroides infection as well as for epidemiological purposes.

Host-parasite association of Placobdella costata (Glossiphoniidae: Hirudinea) and Mauremys leprosa (Geoemydidae: Testudinoidea) in aquatic ecosystems of Morocco.

Emys orbicularis (Linnaeus, 1758) was considered as a specific host of Placobdella costata (Fr. Mûller, 1846). However, since the parasite was recorded from outside the distribution area of its host, some authors suggested a possible relationship with other hosts. Although two accidental associations were found with another turtle, Mauremys leprosa (Schweigger, 1812), the obtained data remain insufficient to better understand this discovered host-parasite ecological system. In this context, the present study was carried out to evaluate the relationship between the Mediterranean pond turtle, M. leprosa, and the freshwater rhynchobdellid leech, P. costata (Hirudinida: Glossiphoniidae), in aquatic ecosystems of Morocco. During the period from April to June 2018, we found leeches attached to turtles in five out of 30 populations sampled with a prevalence of infection significantly higher in adult than that in juvenile turtles. Moreover, the males are the most infested with 51% of the total, followed by females (33.3%) and juveniles (15.7%). The obtained results indicated that 51 turtles were infested by 139 leeches with a mean intensity of infestation of 4.17 ± 0.47 leeches/turtle (up to 10 leeches/turtle). It was higher in males than that in females in almost all sites. The posterior limbs are the most preferred attachment site, and the body condition of turtles was not affected by the intensity of infestation but it is rather a function of altitude. Our findings proved that M. leprosa-P. costata association is more than accidental and that M. leprosa is rather the main host of P. costata in aquatic ecosystems of Morocco.

Further characterisation of Haemocystidium chelodinae-like Haemoproteidae isolated from the Bellinger River snapping turtle (Myuchelys georgesi).

The Bellinger River snapping turtle (Myuchelys georgesi) is endemic to Australia and is confined to a highly restricted distribution in the Bellinger River in New South Wales. Routine veterinary health examinations of 17 healthy turtles were undertaken, along with the collection and analysis of blood samples, during conservation efforts to save the species following a catastrophic population decline. Microscopy analysis of blood films detected Haemoproteidae parasites that morphologically resembled Haemocystidium chelodinae inside turtle erythrocytes. Of the 17 turtles examined, 16 were positive for infection with H. chelodinae by both light microscopy and PCR. DNA sequencing of a partial fragment of the mitochondrial cytochrome b (cytb) gene and phylogenetic analysis identified two different H. chelodinae-like genotypes. The phylogenetic relationship of H. chelodinae-like to other Haemoproteidae species based on cytb sequences grouped H. chelodinae-like into the reptile clade, but revealed the Haemocystidium genus to be paraphyletic as the clade also contained Haemoproteus, thus supporting a re-naming of Haemoproteus species from reptiles to Haemocystidium species. This study reports for the first time the genetic characterisation of H. chelodinae-like organisms isolated from a new Testudine host species, the Bellinger River snapping turtle. As evidence grows, further research will be necessary to understand the mode of transmission and to investigate whether these parasites are pathogenic to their hosts.

The genetic diversity of blood parasites within the freshwater turtles Mauremys leprosa and Emys orbicularis in Tunisia reveals coinfection with Haemogregarina spp.

Haemogregarina species are apicomplexan blood parasites infecting vertebrates such as fish, lizards, and turtles. Due to the high morphological similarity of the erythrocytic stages infecting host species, it has always been a challenge to identify the true diversity of these parasites. Therefore, taxonomic studies are presently based on the combination of morphological and molecular data. In Tunisia, two species of Haemogregarina have been reported within the freshwater turtle Mauremys leprosa (Geoemydidae) for more than 40 years. Since M. leprosa occurs in the same aquatic environments as Emys orbicularis (Emydidae) in Tunisia, our objectives were to assess parasite diversity and specificity on the basis of both morphological and molecular approaches. The turtles were surveyed and sampled across six aquatic areas of Tunisia. Among the 39 specimens of M. leprosa and seven of E. orbicularis that were trapped and investigated, the presence of haemogregarines was detected in the blood of turtles only at sites where leeches were observed. Three 18S variants were identified, which corresponded to three distinct Haemogregarina species, among which one was identified as Haemogregarina stepanowi. The two other species that were detected are likely new to science. Because we show the occurrence of more than one blood parasite species within a single host specimen, our study provides the first report of coinfection with molecularly distinct Haemogregarina spp.

Absence of Hemolivia mauritanica (Apicomplexa: Haemogregarinidae) in natural populations of Testudo graeca in Morocco.

During spring 2018, we captured 101 spur-thighed tortoises, Testudo graeca, from seven localities in central Morocco. All tortoises were examined for the presence of blood parasites Hemolivia mauritanica and Hyalomma aegyptium ticks, the known vectors. We looked for H. mauritanica infection by examination of blood smears and by genetic analysis with PCR using extractions from both tortoises and ticks. On all tortoises collected, 71.29% were infested with at least one tick, with a mean infestation intensity of 7.12 ticks/tortoise and maximum of 15.55 ticks/tortoises in Had Draa locality (Essaouira region). Although the definitive host is present and abundant in all tortoise populations, all blood samples were Hemolivia-negative. Our results support and confirm the finding of studies previously conducted in other populations of Morocco and indicate that H. mauritanica has a narrower distribution range than its tick vector.

Eye to eye: classification of conjunctival sac polystomes (Monogenea: Polystomatidae) revisited with the description of three new genera Apaloneotrema n. g., Aussietrema n. g. and Fornixtrema n. g.

Polystomes (Monogenea: Polystomatidae) of freshwater turtles are currently represented by five genera, namely Neopolystoma, Polystomoides, Polystomoidella, Uropolystomoides and Uteropolystomoides. These parasites can infect the urinary, oral and/or the conjunctival sac systems of their hosts, showing strict site specificity. A recent phylogenetic study showed that the two most diverse genera within chelonian polystomes, i.e. Neopolystoma and Polystomoides, are not monophyletic. Furthermore, polystomes infecting the conjunctival sacs of their host, except for one species, formed a robust lineage. A fusiform egg shape has been reported for conjunctival sac polystomes and it was assumed that this characteristic could be a good character for the systematics of polystomes. Our objective in the present work was, therefore, to study more in depth the morphology of polystomes collected from the conjunctival sacs of chelonians to find characters defining a putative new genus. To achieve this objective, more specimens were collected in 2018 and 2019 from turtles sampled in North Carolina and Florida (USA) to extend taxon sampling for the phylogenetic analysis. Morphological characters of relevant polystome specimens were re-examined from several collections from Asia, Australia, Europe, South Africa, South America and North America. Based on a Bayesian tree inferred from the analysis of four concatenated genes, namely 12S, 18S, 28S and COI, polystomes found in the conjunctival sacs were grouped in three distinct lineages, the first one including a single species infecting an Australian pleurodire turtle; the second one including eleven species infecting cryptodire turtles of South America, North America and Asia; and the last one including a single species infecting a softshell cryptodire turtle of North America. Based on observations of live specimens by Dr. Sylvie Pichelin and our morphological analysis, the conjunctival sac polystomes from Australian turtles are small, cannot extend their body significantly, have a spherical ovary and egg, have a large genital bulb and possess latero-ventral vaginae at the level of the testis. Based on observations of live specimens and morphological analysis of whole mounted specimens, polystomes of the second lineage share the following morphological characteristics: the ability to stretch out and double their length, a long oval ovary, a separate egg-cell-maturation-chamber, fusiform to diamond-shaped eggs with acute tips, small genital bulb and vaginae peripheral on the side of the body at the level of the testis. The polystome species of the third lineage occupies a basal position, has the ability to stretch out and possess an elongated ovary, a large fusiform egg with rounded tips, a small genital bulb and small latero-ventral vaginae at the level of the ovary. These three distinct conjunctival sac polystome lineages are herein described as separate new genera, Aussietrema, Fornixtrema and Apaloneotrema, respectively.

A new species and emendation of the seldom reported Enterohaematotrema Mehra, 1940 (Digenea: Schistosomatoidea), including a revised phylogenetic hypothesis for turtle blood flukes.

Enterohaematotrema Mehra, 1940 is emended herein based upon a review of the literature and a description of a new species (Enterohaematotrema triettruongi n. sp.) infecting yellow-headed temple turtles, Heosemys annandalii (Boulenger) (Cryptodira: Geoemydidae), in the Mekong River, Vietnam. The new species differs from the published descriptions of its congeners Enterohaematotrema palaeorticum Mehra, 1940 and Enterohaematotrema hepaticum (Simha, 1958) Simha & Chattopadhyaya, 1980 by having two distinctive oesophageal glands, a short and eversible cirrus (vs protrusive with 3 distinct processes), a dorsal common genital pore that is sinistral (vs ventral and medial), a transverse (vs longitudinal) external seminal vesicle, an oviducal seminal receptacle that is sinistral (vs dextral), and a vitellarium distributing from the caecal bifurcation (anterior to the ventral sucker) to the caecal tips (vs vitellarium not extending anteriad beyond ventral sucker in E. palaeorticum or vitellarium wholly posterior to the terminal genitalia in E. hepaticum). A phylogenetic analysis of the D1-D3 domains of the nuclear large subunit ribosomal DNA (28S) recovered Enterohaematotrema and Platt Roberts & Bullard, 2016 as sister taxa that share a recent common ancestor with the clade comprising Ruavermis Dutton & Bullard, 2020 and Coeuritrema Mehra, 1933. These flukes collectively comprise a monophyletic group of southeast Asian turtle blood flukes. This analysis also indicated that the massive, longitudinal metraterm of species of Enterohaematotrema and Uterotrema Platt & Pichelin, 1994 represents homoplasy (convergent evolution). The present study comprises the first morphological study of original specimens of any species of Enterohematotrema in more than 50 years and is the first molecular phylogenetic placement of the genus among the various turtle blood fluke lineages.