Larger Common River Frogs (Amietia delalandii) have Fewer and Shorter Tissue Microplastic Fibres than Smaller Frogs.

Little is known about microplastics (MPs) in adult frogs. We investigated MPs in adult Common River Frogs (Amietia delalandii) from Potchefstroom, South Africa. Five kinds of samples were analysed: natural water, water used to rinse the skin, skin, intestine, and the remainder of the body (corpus). Tissues were digested. Microplastics occurred in all frogs and sample types (1128 MPs counted). Fibres were the most prevalent MP. Fibre lengths were between 28 and 4300 µm, either polyester or polyvinyl alcohol. MPs in skin were likely derived from the ambient, and MPs in the corpus from translocation via the skin. Fibres in tissues were significantly shorter in larger frogs, a phenomenon we provisionally assign to in situ biodegradation. Microplastics in frogs can potentially be transferred through the food web to higher trophic levels. This study provides the first evidence of MPs in adult frog tissues and avenues for further investigations.

Taxonomic re-evaluation of African anuran trypanosomes with the redescription and molecular diagnosis of Trypanosoma (Trypanosoma) nelspruitense Laveran, 1904 and Trypanosoma (Haematomonas) grandicolor Pienaar, 1962.

The aquatic and terrestrial clades of species of Trypanosoma could provide insight into the evolutionary history of the genus, as well as complementary information for biomedical studies of medically and economically important species of Trypanosoma. The ecological interactions and phylogeny of aquatic trypanosomes are currently not well-understood, mostly due to their complex life cycles and a deficiency of data. The species of Trypanosoma from African anuran hosts are of the least understood taxa in the genus. Trypanosomes were collected from South African frogs and subjected to morphological and phylogenetic analyses. This study redescribes Trypanosoma (Trypanosoma) nelspruitense Laveran, 1904 and Trypanosoma (Haematomonas) grandicolor Pienaar, 1962, with morphological and molecular data. The present study aims to create a platform for further future research on African anuran trypanosomes.

Infection patterns of helminth community in black rockcod Notothenia coriiceps in West Antarctica over a 6-year term.

Patterns of the rockcod Notothenia coriiceps infection with helminths were analysed to understand the dynamics of parasite communities in this Antarctic fish and to test their stability over time. The study was performed using helminth samples collected from 183 N. coriiceps in 2014-2015 and 2020-2021 in the vicinity of the Ukrainian Antarctic station (UAS) "Akademik Vernadsky", Galindez Island, Argentine Islands, West Antarctica. Overall, 25 helminth taxonomical categories (nine trematodes, four cestodes, five nematodes, and seven acanthocephalans) were subjected to analysis. A direct comparison of the helminth population characteristics showed that nine species significantly changed their infection parameters during the 6 years between the samples. Seven of them (Pseudoterranova sp., Contracaecum sp., Ascarophis nototheniae, monolocular metacestodes, bilocular metacestodes, Metacanthocephalus rennicki, and Diphyllobothrium sp.) were found to have a significant impact on the differences between helminth infracommunities in 2014-2015 and 2020-2021. Most studied patterns of helminth component community appeared to show a stable tendency, and observed fluctuations were close to the steady trend. Slight but significant changes in the infection patterns observed in this study might have been caused by changes in the populations of intermediate, paratenic, and definitive hosts of helminths (marine invertebrates, mammals, and birds), which participate in helminth transmission in Antarctic ecosystems.

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.

Polystoma luohetong n. sp. (Monogenea: Polystomatidae) from Rana chaochiaoensis Liu (Amphibia: Ranidae) in China.

Polystoma chaochiaoensis from the urinary bladder of the chaochiao frog Rana chaochiaoensis Liu was briefly described in a symposium abstract and presented at the Third Symposium on Parasitology of China in 1990. Types were not assigned and the original specimens collected are no longer available. The morphological description was incomplete and no illustrations were provided. We consider Polystoma chaochiaoensis a nomen nudum and provide a full description for the species and assign types. Based on morphological characteristics and molecular data of partial 18S rDNA sequences, we describe this species as Polystoma luohetong n. sp. Out of 578 frogs examined, 16 male and 38 female frogs were infected (prevalence 9.3%; mean intensity 1.02). Polystoma luohetong n. sp. is distinguished from all other Polystoma species by the presense of a prominent crest on the hamulus as well as by the shape and size of marginal hooklets and the intestinal arrangement. Furthermore, the phylogentic analysis based on the 18S rRNA gene shows Polystoma luohetong n. sp. well nested within the Ploystoma clade and as a sister taxon to Polystoma integerrimum.

Monophyly of the species of Hepatozoon (Adeleorina: Hepatozoidae) parasitizing (African) anurans, with the description of three new species from hyperoliid frogs in South Africa.

Haemogregarines (Apicomplexa: Adeleiorina) are a diverse group of haemoparasites reported from almost all vertebrate classes. The most commonly recorded haemogregarines to parasitize anurans are species of Hepatozoon Miller, 1908. To date 16 Hepatozoon species have been described from anurans in Africa, with only a single species, Hepatozoon hyperolli (Hoare, 1932), infecting a member of the Hyperoliidae. Furthermore, only two Hepatozoon species are known from South African anurans, namely Hepatozoon theileri (Laveran, 1905) and Hepatozoon ixoxo Netherlands, Cook and Smit, 2014, from Amietia delalandii (syn. Amietia quecketti) and three Sclerophrys species, respectively. Blood samples were collected from a total of 225 individuals representing nine hyperoliid species from several localities throughout northern KwaZulu-Natal, South Africa. Twenty frogs from three species were found positive for haemogregarines, namely Afrixalus fornasinii (6/14), Hyperolius argus (2/39), and Hyperolius marmoratus (12/74). Based on morphological characteristics, morphometrics and molecular findings three new haemogregarine species, Hepatozoon involucrum Netherlands, Cook and Smit n. sp., Hepatozoon tenuis Netherlands, Cook and Smit n. sp. and Hepatozoon thori Netherlands, Cook and Smit n. sp., are described from hyperoliid hosts. Furthermore, molecular analyses show anuran Hepatozoon species to be a separate monophyletic group, with species isolated from African hosts forming a monophyletic clade within this cluster.

Linking organochlorine exposure to biomarker response patterns in Anurans: a case study of Müller's clawed frog (Xenopus muelleri) from a tropical malaria vector control region.

Organochlorine pesticides are highly persistent in aquatic ecosystems. Amphibians, specifically anurans, play an intricate part in the aquatic food web, and have very permeable skin which makes them prone to bioaccumulation of persistent pollutants. In this study the bioaccumulation of various legacy organochlorine pesticides (OCPs)-including dichlorodiphenyltrichloroethane (DDT), currently used for malaria vector control (MVC)-was assessed along with a set of biomarker responses in Müller's clawed frog Xenopus muelleri collected from the lower Phongolo River floodplain in South Africa. Possible relationships between bioaccumulation and biomarkers (of exposure, oxidative stress biomarkers, and cellular energy allocation) alongside their temporal changes were investigated. The OCP concentrations showed a significant increase over time for the duration of the study. The increase correlated negatively with rainfall from the region. DDT levels were well below expected effects levels with p,p-DDE being the main contributing metabolite. The results of this study indicate OCPs actively accumulate at sub-lethal levels in aquatic frogs from the study area, while showing possible relations towards some of the biochemical stress responses measured. Most notable were negative relationships indicated between p,p-DDE and acetylcholinesterase, malondialdehyde, and carbohydrates and protein energy availability. Levels of DDT were not found to be significantly higher than other legacy pesticides in the frog tissue, although evidence of newly introduced DDT in the frog tissue was found. Further investigation about sub-lethal effects of these pesticides on anurans is required to gain better insight into their full impact on animal livelihood.

New information on morphology and molecular data of camallanid nematodes parasitising Xenopus laevis (Anura: Pipidae) in South Africa.

Three species of nematodes from the Camallanidae that are known to infect Xenopus laevis Daudin (Anura: Pipidae) were collected from several localities across South Africa. New data on morphology, partial 28S and cox1 genes, infection levels and distribution are presented herein. The most common species, Batrachocamallanus slomei Southwell et Kirshner, 1937, from the stomach and less often oesophagus, was found in eight localities. Camallanus kaapstaadi Southwell et Kirshner, 1937, also from the oesophagus, was found in two localities and C. xenopodis Jackson et Tinsley, 1995, from the intestine, at a single locality. New localities for both C. kaapstaadi and C. xenopodis provide a geographical range extension. Males of C. xenopodis are described for the first time herein. The existence of a left spicule in the males of both the species of Camallanus Railliet and Henry, 1915 is confirmed and measurements are provided. Although C. xenopodis is distinguished from C. mazabukae Kung, 1948 in the present study, we suggest greater sampling effort in other African amphibians to confirm the species status of the latter taxon. Finally, the new molecular data showed distant relationships between collected species of Camallanus and species parasitising fish and freshwater turtles.

Ultrastructural Comparison of Hepatozoon ixoxo and Hepatozoon theileri (Adeleorina: Hepatozoidae), Parasitising South African Anurans.

To date, only two haemogregarine parasite species have been described from South African anurans: Hepatozoon ixoxo, infecting toads of the genus Sclerophrys (syn. Amietophrynus); and Hepatozoon theileri, parasitising the common river frog, Amietia quecketti. Both species have been characterised using limited morphology, and molecular data from PCR amplified fragments of the 18S rRNA gene. However, no ultrastructural work has been performed thus far. The aim of this study was to add descriptive information on the two species by studying their ultrastructural morphology. Mature gamont stages, common in the peripheral blood of infected frogs, were examined by transmission electron microscopy. Results indicate that H. ixoxo and H. theileri share typical apicomplexan characteristics, but differ markedly in their external cellular structure. Hepatozoon ixoxo is an encapsulated parasite presenting a prominent cap at the truncate pole, and shows no visible modifications to the host cell membrane. In comparison, H. theileri does not present a capsule or cap, and produces marked morphological changes to its host cell. Scanning electron microscopy was performed to further examine the cytopathological effects of H. theileri, and results revealed small, knob-like protrusions on the erythrocyte surface, as well as notable distortion of the overall shape of the host cell.

Evolutionary processes involved in the diversification of chelonian and mammal polystomatid parasites (Platyhelminthes, Monogenea, Polystomatidae) revealed by palaeoecology of their hosts.

Polystomatid flatworms (Platyhelminthes) are monogenean parasites that infect exclusively aquatic or semi-aquatic sarcopterygians such as the Australian lungfish, amphibians, freshwater turtles and the African common hippopotamus. Previous studies on the phylogenetic relationships of these parasites, excluding Oculotrema hippopotami infecting common hippos, showed a global coevolution between hosts and their parasites at a macroevolutionary scale. These studies also demonstrated a strong correlation between the diversification of early neobatrachian polystomes and Gondwana breakup in the Mesozoic period. However the origin of chelonian polystomes is still in question as a switch from presumably primitive aquatic amniotes to turtles at the time of their first appearance, or soon after during their radiation, was assumed. In order to resolve this sticking point, we extended the phylogeny of polystomes with broader parasite sampling, i.e. 55 polystome species including Nanopolystoma tinsleyi a polystome infecting caecilians and O. hippopotami, and larger set of sequence data covering two nuclear and two mitochondrial genes coding for the ribosomal RNA 18S and 28S, the Cytochrome c Oxidase I and the ribosomal RNA 12S, respectively. The secondary structure of nuclear rRNAs genes (stems and loops) was taken into account for sequence alignments and Bayesian analyses were performed based on the appropriate models of evolution selected independently for the four designed partitions. Molecular calibrations were also conducted for dating the main speciation events in the polystome tree. The phylogenetic position of chelonian parasites that are phylogenetically closer to N. tinsleyi than all other amphibian polystomes and molecular time estimates suggest that these parasites originated following a switch from caecilians, at a geological period when primitive turtles may already have adapted to an aquatic life style, i.e. at about 178Million years ago, or a little later when the crown group of extant turtles have already diversified, i.e. at about 152Mya. Similarly, because O. hippopotami constitutes the sister group of chelonian parasites, proposing that an African caecilian could be the ancestral host for this polystome species seems at this stage the most likely hypothesis to explain its occurrence within the common hippo. Regardless of the scenario that may be predicted to explain the origin of polystomes within aquatic or semi-aquatic amniotes, their presence and evolution are indicative of early aquatic ecological habits within ancestral lineages.

Some nematodes of the genus Rhabdias Stiles et Hassall, 1905 (Nematoda: Rhabdiasidae) parasitising amphibians in French Guiana.

Based on material collected from Rhinella cf. margaritifera (Laurenti) and Rhi. marina (Linnaeus) (Anura: Bufonidae) during a parasite survey of the herpetofauna of French Guiana, updated descriptions of Rhabdias androgyna Kloss, 1971 and R. fuelleborni Travassos, 1926 are presented. In addition to metrical data, which may overlap in closely related species, emphasis is placed on qualitative characters. Rhabdias androgyna is distinguished by the unique presence of an outer and inner cephalic cuticular inflation, a shoulder-like broadening of the body at the anterior end, a wide and shallow buccal capsule (average buccal ratio 0.36) with serrated lumen in apical view, a prominent anterior dilatation of the oesophagus, and the presence of an additional posterior dilatation anterior to the oesophageal bulb. Characters that may help to differentiate R. fuelleborni from closely related species parasitising the Rhi. marina species group are the presence of six relatively uniform lips, and the division of the buccal capsule into an anterior and posterior segment, with differently structured walls. Both the presence of R. androgyna and R. fuelleborni in French Guiana constitute new geographic records. A single specimen of Rhabdias sp. is described from Pristimantis chiastonotus (Lynch et Hoegmood) (Anura: Craugastoridae). This species differs from all its Neotropical congeners by the distinct globular swelling of its head, similar to that seen in only one Palaearctic and one Afrotropical Rhabdias species. A list of species of Rhabdias parasitising amphibians in the Netropical Realm is also provided.

Multiple emergences of genetically diverse amphibian-infecting chytrids include a globalized hypervirulent recombinant lineage.

Batrachochytrium dendrobatidis (Bd) is a globally ubiquitous fungal infection that has emerged to become a primary driver of amphibian biodiversity loss. Despite widespread effort to understand the emergence of this panzootic, the origins of the infection, its patterns of global spread, and principle mode of evolution remain largely unknown. Using comparative population genomics, we discovered three deeply diverged lineages of Bd associated with amphibians. Two of these lineages were found in multiple continents and are associated with known introductions by the amphibian trade. We found that isolates belonging to one clade, the global panzootic lineage (BdGPL) have emerged across at least five continents during the 20th century and are associated with the onset of epizootics in North America, Central America, the Caribbean, Australia, and Europe. The two newly identified divergent lineages, Cape lineage (BdCAPE) and Swiss lineage (BdCH), were found to differ in morphological traits when compared against one another and BdGPL, and we show that BdGPL is hypervirulent. BdGPL uniquely bears the hallmarks of genomic recombination, manifested as extensive intergenomic phylogenetic conflict and patchily distributed heterozygosity. We postulate that contact between previously genetically isolated allopatric populations of Bd may have allowed recombination to occur, resulting in the generation, spread, and invasion of the hypervirulent BdGPL leading to contemporary disease-driven losses in amphibian biodiversity.

Assessment of platyhelminth diversity within amphibians of French Guiana revealed a new species of Nanopolystoma (Monogenea: Polystomatidae) in the caecilian Typhlonectes compressicauda.

An expedition was undertaken to French Guiana in search of amphibian parasites. Of the 23 anuran species collected and screened for polystomes, the toad Rhinella margaritifera (Laurenti) was the sole species found to be infected with a polystome, namely Wetapolystoma almae Gray, 1983. Of the two caecilian species collected, a new species of Nanopolystoma du Preez, Huyse et Wilkinson, 2008 was discovered from the urinary bladder of the aquatic caecilian Typhlonectes compressicauda (Duméril et Bibron). The small size of the mature worm, two non-diverticulated caeca of equal length that are non-confluent posteriorly, vitelline follicles in two dense lateral fields, a single follicular testis in the middle of the body, small ovary and a single operculated egg in utero, vaginae present and the caecilian host allowed the identification of the specimen as Nanopolystoma. Larger body size, hamulus length, egg diameter and occurrence in the caecilian family Typhlonectidae distinguishes the new species from the two other known polystomes in Nanopolystoma; thus, the description of Nanopolystoma tinsleyi sp. n. is provided within this paper.

Redescription and molecular diagnosis of Hepatozoon theileri (Laveran, 1905) (Apicomplexa: Adeleorina: Hepatozoidae), infecting Amietia quecketti (Anura: Pyxicephalidae).

Blood smears prepared from the peripheral blood of 20 wild caught Amietia quecketti (Boulenger) from the North-West University Botanical Gardens, North West Province, South Africa, were examined for the presence of haemogregarines. A haemogregarine species comparative in morphology, host and geographical locality to that of Haemogregarina theileri Laveran, 1905 was detected. The original description of H. theileri was based solely on frog peripheral blood gamont stages. Later, further parasite stages, including trophozoites and merogonic liver stages, were recorded in a related Amietia sp. from equatorial Africa. This species was originally classified as a member of the genus Haemogregarina Danilewsky, 1885, but due to the close life cycle and morphological resemblance to those of Hepatozoon species, H. theileri was later transferred from Haemogregarina to Hepatozoon Miller, 1908. In the present study, meront and merozoite stages not described before, along with previously observed trophozoite, immature and mature gamont stages, are described from the peripheral blood of hosts. In addition, comparative phylogenetic analysis of the partial 18S rDNA sequence of Hepatozoon theileri to those of other haemogregarine species, including those of species of Hepatozoon and a Haemogregarina, support the taxonomic transfer of H. theileri to Hepatozoon, nesting H. theileri within a clade comprising species parasitising other amphibians. This is the first molecular and phylogenetic analysis of an African anuran species of Hepatozoon.

Barcoding and traditional health practitioner perspectives are informative to monitor and conserve frogs and reptiles traded for traditional medicine in urban South Africa.

Previous literature suggests that Indigenous cultural practices, specifically traditional medicine, are commonplace among urban communities contrary to the general conception that such practices are restricted to rural societies. We reviewed previous literature for records of herptiles (frog and reptile species) sold by traditional health practitioners in urban South Africa, then used visual confirmation surveys, DNA barcoding and folk taxonomy to identify the herptile species that were on sale. Additionally, we interviewed 11 IsiZulu and SePedi speaking traditional health practitioners to document details of the collection and pricing of herptile specimens along with the practitioners' views of current conservation measures for traditional medicine markets. The 34 herptile species recorded in previous literature on traditional medicine markets included endangered and non-native species. Spectrophotometry measurements of the DNA we extracted from the tissue of herptiles used in traditional medicine were an unreliable predictor of whether those extractions would be suitable for further experimental work. From our initial set of 111 tissue samples, 81 sequencing reactions were successful and 55 of those sequences had species-level matches to COI reference sequences on the NCBI GenBank and/or BOLD databases. Molecular identification revealed that traditional health practitioners correctly labelled 77% of the samples that we successfully identified with DNA barcoding in this study. Our mixed methodology approach is useful for conservation planning as it updates knowledge of animal use in Indigenous remedies and can accurately identify species of high conservation priority. Furthermore, this study highlights the possibility of collaborative conservation planning with traditional health practitioners.

Correlating early evolution of parasitic platyhelminths to Gondwana breakup.

Investigating patterns and processes of parasite diversification over ancient geological periods should involve comparisons of host and parasite phylogenies in a biogeographic context. It has been shown previously that the geographical distribution of host-specific parasites of sarcopterygians was guided, from Palaeozoic to Cainozoic times, mostly by evolution and diversification of their freshwater hosts. Here, we propose phylogenies of neobatrachian frogs and their specific parasites (Platyhelminthes, Monogenea) to investigate coevolutionary processes and historical biogeography of polystomes and further discuss all the possible assumptions that may account for the early evolution of these parasites. Phylogenetic analyses of concatenated rRNA nuclear genes (18S and partial 28S) supplemented by cophylogenetic and biogeographic vicariance analyses reveal four main parasite lineages that can be ascribed to centers of diversity, namely Australia, India, Africa, and South America. In addition, the relationships among these biogeographical monophyletic groups, substantiated by molecular dating, reflect sequential origins during the breakup of Gondwana. The Australian polystome lineage may have been isolated during the first stages of the breakup, whereas the Indian lineage would have arisen after the complete separation of western and eastern Gondwanan components. Next, polystomes would have codiverged with hyloid sensu stricto and ranoid frog lineages before the completion of South American and African plate separation. Ultimately, they would have undergone an extensive diversification in South America when their ancestral host families diversified. Therefore, the presence of polystome parasites in specific anuran host clades and in discrete geographic areas reveals the importance of biogeographic vicariance in diversification processes and supports the occurrence and radiation of amphibians over ancient and recent geological periods.

ParaSiteDB - A user-friendly database for managing a parasitological collection.

ParaSiteDB is an application for arranging and managing a parasitological collection. It has been designed to provide a user-friendly, easily manageable and searchable site and is suitable for small to bigger collections. The source code of the application is available on GitHub: https://github.com/goobar4/aacrg. The demonstration version of the application is available on https://syrota.info/wormbasehttp://syrota.info/wormbase.

Two new species of Neofoleyellides (Nematoda: Onchocercidae) parasitising anuran amphibians in South Africa.

The genus Neofoleyellides was recently erected for a single species, Neofoleyellides boerewors from bufonid hosts in South Africa. In present study, we discovered two undescribed species of Neofoleyellides, namely N. steyni n. sp. and N. martini n. sp. parasitising frogs Amietia delalandii and Leptopelis natalensis, respectively. Both species differ from N. boerewors and between each other in shape and relative length of oesophagus, size of spicules, arrangement of genital papillae and morphology of caudal alae in males. Phylogenetic analysis based on concatenated fragments of the 18S ribosomal ribonucleic acid (18S rRNA) and the Cytochrome oxidase c subunit I (COI) genes confirmed both species as Neofoleyellides sister to Icosiellinae and Oswaldofilariinae.

First Record of a Polystome from Alligator Snapping Turtle, Macrochelys temminckii (Cryptodira: Chelydridae) or Mississippi; with Comments on "Neopolystoma orbiculare (Stunkard, 1916)" and Its Junior Subjective Synonyms.

Herein, we describe several newly-collected specimens of Neopolystoma cf. orbiculare from the urinary bladder of 2 alligator snapping turtles, Macrochelys temminckii (Troost in Harland, 1835) (Cryptodira: Chelydridae Gray, 1831) from Comet Lake (30°35'46.94″N, 88°36'3.12″W), Pascagoula River, Mississippi. Our specimens differed from all previous descriptions of N. orbiculare and its junior subjective synonyms by the combination of having intestinal ceca adorned with triangular pockets and that terminate dorsal to the haptor, distinctive hooklets each having a handle and guard of approximately equal length and having a much longer and curved blade, 16 genital coronet spines that each possess 1-2 flanges per spine, pre-testicular vaginal pores, and vaginal ducts that are anterior to the junction of the oviduct and genito-intestinal canal. Some of our specimens were enantiomorphic (4 and 3 had a dextral and sinistral ovary, respectively). Nucleotide sequences (large subunit ribosomal DNA [28S], small subunit ribosomal DNA [18S], and cytochrome oxidase subunit 1 mitochondrial gene [COI]) for our specimens were most similar to GenBank sequences ascribed to N. orbiculare. Single-gene and concatenated phylogenetic analyses confirmed that NeopolystomaPrice, 1939 is polyphyletic and that our isolates share a recent common ancestor with those ascribed to N. orbiculare. This is the first record of a polystomatid from Mississippi, from the Pascagoula River, and from the alligator snapping turtle (and only the second species of Neopolystoma reported from any snapping turtle).

Molecular confirmation of high prevalence of species of Hepatozoon infection in free-ranging African wild dogs (Lycaon pictus) in the Kruger National Park, South Africa.

Reports in the literature indicate that species of Hepatozoon commonly occur in African wild dog (AWD) or painted wolf (Lycaon pictus) populations. These findings were based on examination of blood smears by microscopy, and specific identity of the Hepatozoon sp. gamonts seen could not be confirmed. We present the first in-depth molecular data on the prevalence of species of Hepatozoon in a free-ranging AWD population. In a general health survey of AWDs in the Kruger National Park, blood specimens (n = 74) collected from 54 individuals were examined for the presence of Hepatozoon spp. At first sampling, specimens from 42 of 54 individuals (77.7%) were positive, based on the primer set HepF300 and HepR900. Twenty individuals were resampled between 51 and 69 days after first sampling; one of these was resampled twice. Samples from six individuals that had tested negative previously now reacted positive. Assuming that all 54 individuals were still alive, the prevalence had therefore increased to 48 individuals infected, or 88.8%. Resultant 18S rDNA sequences isolated from these specimens share high similarity to other Hepatozoon canis genotypes. Phylogenetic analysis recovered the Hepatozoon sp. isolated from AWDs within the H. canis cluster, which includes species of Hepatozoon from other canid and tick hosts.