First record of Sebekia oxycephala (Pentastomida: Sebekidae) infecting Helicops angulatus (Reptilia: Colubridae) in Northeast Brazil.

In this study, we investigated the infection by pulmonary parasites in the water snake Helicops angulatus. We found four nymphs of the pentastomide parasite Sebekia oxycephala, parasitizing two adult females H. angulatus collected in the municipality of Rio Tinto, Paraíba, Brazil. This occurrence is likely due to the nymphs of this parasite directly infecting aquatic intermediate hosts, such as fish. The result of this study represents the first record of pentastomid infecting H. angulatus.

A 50-year-old redescription: molecular and morphometric characterization of Hepatozoon carinicauda Pessôa and Cavalheiro, 1969 in the brown-banded water snake Helicops angulatus (Linnaeus, 1758).

The combined use of molecular and microscopic techniques has become an increasingly common and efficient practice for the taxonomic and evolutionary understanding of single-celled parasites such as haemogregarines. Based on this integrative approach, we characterized Hepatozoon found in Helicops angulatus snakes from the Eastern Amazonia, Brazil. The gamonts observed caused cell hypertrophy and were extremely elongated and, in some cases, piriform (mean dimensions: 25.3 ± 1.9 × 8.6 ± 1.3 µm). These morphological features correspond to Hepatozoon carinicauda, described 50 years ago in the snake Helicops carinicaudus in the southeast region of Brazil. Phylogenetic and genetic divergence analyses, performed with the sequence obtained from the amplification of a 590 bp fragment of the 18S rRNA gene, revealed that Hepatozoon in He. angulatus differed from the other lineages retrieved from GenBank, and was clustered singly in both the phylogenetic tree and the haplotype network. The integration of these data allowed the identification of H. carinicauda in a new aquatic host, and increased the knowledge of its geographical distribution. Therefore, the present study included the first redescription of a Hepatozoon species in a snake from the Brazilian Amazon.

Sarcocystis sp. infection (Apicomplexa: Sarcocystidae) in invasive California kingsnake Lampropeltis californiae (Serpentes: Colubridae) in Gran Canaria.

Invasive species pose a threat not only to biodiversity because they displace or compete with native fauna, but also because of the pathogens they can host. The Canary Islands are an Atlantic biodiversity hotspot threatened by increasing numbers of invasive species, including the California kingsnake Lampropeltis californiae, which was recently introduced to Gran Canaria. Seventy-seven snakes were examined for gastrointestinal parasites in 2019­2020. Sporocysts of Sarcocystis sp. were detected in 10 of them; detection of gamogonia stages in histological sections of 3 snakes confirmed the snake as a definitive host. Partial ssrDNA was amplified using SarcoFext/SarcoRext primers; an additional sequence of Sarcocystis was obtained from the tail muscle of the endemic Gran Canaria giant lizard Gallotia stehlini for a comparison. Identical ssrDNA sequences of unknown Sarcocystis sp. were obtained from 5 different snakes. Phylogenetic analysis showed that Sarcocystis sp. isolated from invasive California kingsnakes is unrelated to Sarcocystis provisionally considered S. stehlini from the endemic lizard. The dixenous coccidia are rarely reported to invade new predator­prey systems. However, the present data suggest that previously unknown Sarcocystis sp. is circulating among invasive snakes and as yet unknown vertebrate intermediate hosts, with undetermined consequences for the Gran Canaria ecosystem.

Hepatozoon cevapii (Apicomplexa: Hepatozoidae) in the Thamnodynastes lanei snake (Colubridae, Tachymenini) from the Eastern Amazon, Brazil.

The aim of the present study was to verify the occurrence of hemogregarines in the colubrid snake Thamnodynastes lanei from the eastern Amazon region of Brazil. Intraerythrocytic gamonts with mean dimensions of 14.8 ± 1.8 × 4.0 ± 0.7 µm and encapsulated gamonts with mean dimensions of 15.3 ± 1.1 × 4.8 ± 0.5 µm were observed. Through morphological and molecular data based on the partial 18S rDNA gene, the parasite was identified as Hepatozoon cevapii, originally described in the viperid snake Crotalus durissus terrificus from the southeast region of Brazil. Thus, the findings of the present study extend the geographic range of H. cevapii and provide novel Hepatozoon-snake associations.

Parasitological findings in the invasive California kingsnake (Lampropeltis californiae) in Gran Canaria, Spain.

The California kingsnake (Lampropeltis californiae), native to North America, is a significant threat to the conservation of endemic species in the Spanish Macaronesian island of Gran Canaria. However, its role disseminating potential invasive parasites, such as zoonotic pentastomids, has not been proven. Among its parasitic fauna, only protistans have been documented, in contrast to other Lampropeltis spp., which are known to carry pentastomids. Thus, a parasitological study is urgently required. Between 2016 and 2018, a total of 108 snakes were necropsied and stool samples examined. A single snake was infested with Ophionyssus natricis, and another individual with Serpentirhabdias sp. Only this latter snake presented gross lesions, characterized by granulomatous pneumonia. No Pentastomida were found. By contrast, almost the entire population (98.5%) was infested with larval helminths (three different nematode and two cestode species), characterized by granulomatous gastrointestinal serositis. This suggests the snake poses a 'dead end' host for local parasites. Based on these findings, snakes in Gran Canaria carry potential zoonotic mites, which along with Serpentirhabdias sp. could represent a threat to endemic lizards. The presence of metazoan parasites and their lesions are reported for the first time in the California kingsnake.

The mitogenome of Ophidascaris wangi isolated from snakes in China.

Different species of the genus Ophidascaris (Baylis, 1921; Nematoda: Ascaridida, Ascaridoidea) are intestinal parasites of various snake species. More than 30 Ophidascaris species have been reported worldwide; however, few molecular genetic studies have been conducted on this genus. We sequenced the complete mitogenome of Ophidascaris wangi parasitizing two snake species of the family Colubridae, i.e., Elaphe carinata (Günther, 1864) and Dinodon rufozonatum. The mitogenome sequence of O. wangi was approximately 14,660 base pairs (bp) long and encoded 36 genes, including 12 protein-coding genes (PCGs), 2 ribosomal RNA (rRNA) genes, and 22 transfer RNA genes. Gene arrangement, genome content, and transcription direction were in line with those in Toxascaris leonina (Linstow, 1902; Ascaridida: Ascarididae). Phylogenetics of O. wangi and other ascaridoids were reconstructed based on the concatenated amino acid sequences of 12 PCGs, and on nucleotide sequences of 12 PCGs and two rRNA genes. Phylogenetic analyses were performed using maximum likelihood and Bayesian inference methods, and the results suggested that O. wangi constitutes a sister clade of Ascaris, Parascaris, Baylisascaris, and Toxascaris within the family Ascarididae, which is a sister clade of Toxocaridae. The mitogenome sequence of O. wangi obtained from the present study will be useful for future identification of the nematode worms in the genus Ophidascaris and will increase the understanding of population genetics, molecular epidemiology, and phylogenetics of ascaridoid nematodes in snakes.

Phylogenetic analysis of Hepatozoon spp. (Apicomplexa: Hepatozoidae) infecting Philodryas patagoniensis (Serpentes: Dipsadidae) in Uruguay.

The purpose of this study was to perform a phylogenetic analysis of Hepatozoon spp. infecting Philodryas patagoniensis in Uruguay. Twenty-five road-killed specimens of P. patagoniensis from ten departments were obtained. Samples of blood and/or heart tissue were taken. Polymerase chain reaction (PCR) assay was carried out amplifying a specific target region of the 18S rRNA gene of Hepatozoon spp. Eighteen out of twenty-five samples were positive to Hepatozoon spp., which gave an overall prevalence of 72%. Phylogenetic analyses with the obtained sequences were carried out to determine the relationship with closely related species found in the region. The results revealed that samples were split into two clades with a high bootstrap support. Clade I was formed with Hepatozoon spp. sequences obtained in this study from P. patagoniensis, Hepatozoon cuestensis from Crotalus durissus terrificus and Hepatozoon musa from Philodryas nattereri, and Hepatozoon spp. retrieved from Cerdocyon thous, Hemidactylus mabouia, and Phyllopezus pollicaris from Brazil, respectively. Clade II was grouped with Hepatozoon cevapii and Hepatozoon massardii, both species described for C. d. terrificus from Brazil. This is the first report of Hepatozoon spp. in snakes from Uruguay.

Description, molecular characterization and life cycle of Serpentirhabdias mussuranae n. sp. (Nematoda: Rhabdiasidae) from Clelia clelia (Reptilia: Colubroidea) in Brazil.

Serpentirhabdias mussuranae n. sp. is described from the lungs of the mussurana, Clelia clelia (Daudin, 1803), from vicinities of Lábrea, Amazonas State, Brazil. The species is characterized by the triangular oral opening, the presence of teeth (onchia) in the oesophastome, the excretory glands longer than the oesophagus and the tail abruptly narrowing in its anterior half and gradually tapering in posterior half. Among the Neotropical representatives of the genus, three species are known to possess the onchia in the oesophastome: S. atroxi, S. moi and S. viperidicus. Serpentirhabdias mussuranae n. sp. differs from S. atroxi and S. viperidicus by its triangular shape of the oral opening and the oesophastome in apical view, vs. round in the latter two congeners. Additionally, S. viperidicus has a larger oesophastome, 13-22 micrometers wide and 13-23 micrometers deep. The new species has relatively longer excretory glands than S. moi. The new species is morphologically and genetically close to S. atroxi, S. moi and S. viperidicus, all parasitic in Brazilian snakes, based on the presence of onchia and the comparison of nucleotide sequences of nuclear ribosomal DNA and mitochondrial cox1 gene (differences varied between 3.8% and 7.1%). Data on the life cycle of S. mussuranae n. sp. is provided, and the life cycle is typical of the genus Serpentirhabdias, with the combination of direct development and heterogony. Free-living larval stages and the adults of amphimictic free-living generation are described. The results of molecular phylogenetic analysis based on nuclear ribosomal internal transcribed spacer (ITS) + partial 28S region and partial mitochondrial cox1 gene are provided.

Interrelations among Multiple Metrics of Immune and Physiological Function in a Squamate, the Common Gartersnake (Thamnophis sirtalis).

The field of ecoimmunology has made it clear that individual and ecological contexts are critical for interpreting an animal's immune response. In an effort to better understand the relevance of commonly used immunological assays, we tested how different metrics of immunity and physiological function were interrelated in naturally parasitized individuals of a well-studied reptile, the common gartersnake (Thamnophis sirtalis). Overall, we found that bactericidal ability, an integrative measure of innate immunity, was often correlated with more specific immunological and physiological tests (lysis and oxidative stress) but was not related to tissue-level inflammation that was determined by histopathology. The only hematological metric that correlated with tissue-level inflammation was the prevalence of monocytes in blood smears. Finally, using histological techniques, we describe natural parasitism throughout the organ systems in these individuals, finding that neither the presence nor the burden of parasite load affected the physiological and immune metrics that we measured. By performing comprehensive assessments of physiological and immune processes, we are better able to draw conclusions about how to interpret findings from specific assays in wild organisms.

The first case of genetically confirmed sparganosis (Spirometra erinaceieuropaei) in European reptiles.

Sparganosis is a zoonosis caused by the spargana (larvae) of Spirometra sp. (Diphyllobothriidae). Reptiles are particularly important vectors for the transmission of this parasite in Asia; however, their role in sparganosis spread in European wildlife is unrecognized. We investigated the infection of reptiles with Spirometra sp. in NE Poland, where several mammalian hosts have been identified recently and in the past. Of the 59 dead reptiles, plerocercoids were found in two grass snakes (Natrix natrix) from the Bialowieza Primeval Forest (BPF). The Spirometra erinaceieuropaei species was genetically confirmed using the evolutionary conserved nuclear 18S rRNA gene, and then compared to GenBank deposits. The sequences were identical to previously investigated Spirometra sp. found in Eurasian badger and wild boar from BPF. Our finding is the first genetically confirmed record of Spirometra sp. in reptiles in Europe. Since reptiles are often a component of mammalian diet, they can be a source of Spirometra tapeworm infection in European wildlife; however, further studies are needed to investigate the prevalence of infection in reptiles and other non-mammalian hosts.

Molecular characterisation of Hepatozoon aegypti Bashtar, Boulos & Mehlhorn, 1984 parasitising the blood of Spalerosophis diadema (Serpentes: Colubridae).

Hepatozoon aegypti Bashtar, Boulos & Mehlhorn, 1984 was first described from the blood of the diadem snake (Spalerosophis diadema) in Egypt. During an investigation of the diversity of reptilian haemogregarines in Saudi Arabia, seven diadem snakes (100% of the sample) were found to be highly parasitised by H. aegypti, with an average parasitaemia of 37% per 500 counted erythrocytes. A complete characterisation of this species with morphometrics and 18S rDNA sequence data is therefore presented here. The infection was found to be restricted to the erythrocytes with, frequently, single and, sometimes, double infections. Mature gamonts were sausage-shaped with round posterior and anterior extremities and measured 14 (13-17) × 3.5 (3-5) µm. The infected erythrocytes were hypertrophied with a faintly stained cytoplasm and longitudinally stretched nuclei. The merogonic stages occurred only in the endothelial cells of the snakes' lungs, and no stages were found in other organs. Mature meronts were round in shape, measured 18 (17-21) µm in diameter and were estimated to produce between 9 and 15 merozoites. Phylogenetic analysis based on the partial 18S small subunit ribosomal DNA sequences indicates that Hepatozoon aegypti cluster within a mixed clade of Hepatozoon species parasitising snakes, geckos and rodents from various geographic areas. Our results might reinforce the theory of prey-predator transmission in respect to the relationships of snake-host Hepatozoon species.

A new coccidian parasite (Apicomplexa: Eimeriidae: Eimeria) from the southern black racer, Coluber constrictor priapus (Reptilia: Ophidia: Colubridae) from Arkansas, USA.

Between April 2012 and October 2017, 18 southern black racers, Coluber constrictor priapus, were collected from nine counties of Arkansas (n = 13) and McCurtain County, Oklahoma (n = 5) and their faeces examined for coccidian parasites. One of 18 (6%) C. c. priapus harboured an eimerian that we describe here as new. Oocysts of Eimeria dunni sp. nov. were subspheroidal with a lightly pitted bi-layered wall measuring L × W 24.0 × 21.1 and L/W ratio of 1.2. A micropyle was absent but an oocyst residuum and polar granule were present. Sporocysts were ovoidal and measured 11.9 × 8.1 with L/W of 1.5. A Stieda body was present but substieda and parastieda bodies were absent. The sporocyst residuum was composed of medium-sized granules aligned along perimeter of sporocyst or in a dispersed mass. This represents the first valid eimerian reported from the southern black racer.

Two new species of Hepatozoon (Apicomplexa: Hepatozoidae) parasitising species of Philothamnus (Ophidia: Colubridae) from South Africa.

To date, only a few species of Hepatozoon Miller, 1908 have been described from amphibians and reptiles of South Africa, including two species from anuran hosts, three from saurians, one from chelonians, and two from ophidians. Hepatozoon bitis (Fantham, 1925) and Hepatozoon refringens (Sambon et Seligmann, 1907), parasitising Bitis arientans (Merrem) and Pseudoaspis cana (Linnaeus), respectively, were described in the early 1900s and since then there have been no further species of Hepatozoon described from snakes in South Africa. Blood smears, used in peripheral blood haemogregarine stage morphometrics, and whole blood used in molecular characterisation of haemogregarines were collected from the caudal vein of six snakes of three species, namely Philothamnus hoplogaster (Günther), Philothamnus semivariegatus (Smith) and Philothamnus natalensis natalensis (Smith). For comparison, a comprehensive table summarising available information on species of Hepatozoon from African snakes is presented. Haemogregarines found infecting the snakes from the present study were morphologically and molecularly different from any previously described from Africa and are thus here described as Hepatozoon angeladaviesae sp. n. and Hepatozoon cecilhoarei sp. n. Both haemogregarine species were observed to cause considerable dehaemoglobinisation of the host cell, in case of infection with H. angeladaviesae resulting in a characteristic peripheral undulation of the host cell membrane and karyorrhexis. To the authors' knowledge, these are the first haemogregarines parasitising snakes of the genus Philothamnus Smith described using both morphological and molecular characteristics in Africa.

Sarcocystis pantherophisi n. sp., from Eastern Rat Snakes (Pantherophis alleghaniensis) as Definitive Hosts and Interferon Gamma Gene Knockout Mice as Experimental Intermediate Hosts.

Here, we report a new species, Sarcocystis pantherophisi n. sp., with the Eastern rat snake (Pantherophis alleghaniensis) as natural definitive host and the interferon gamma gene knockout (KO) mouse as the experimental intermediate host. Sporocysts (n = 15) from intestinal contents of the snake were 10.8 × 8.9 µm. Sporocysts were orally infective to KO mice but not to laboratory-raised albino outbred house mice (Mus musculus). The interferon gamma KO mice developed schizont-associated neurological signs, and schizonts were cultivated in vitro from the brain. Mature sarcocysts were found in skeletal muscles of KO mice examined 41 days postinoculation (PI). Sarcocysts were slender, up to 70 µm wide and up to 3.5 mm long. By light microscopy, sarcocysts appeared thin-walled (<1 µm) without projections. By transmission electron microscopy, the sarcocyst wall was a variant of "type 1" (type 1i, new designation). The parasitophorous vacuolar membrane (pvm) had approximately 100-nm-wide × 100-nm-long bleb-like evaginations interspersed with 100-nm-wide × 650-nm-long elongated protrusions at irregular distances, and invaginations into the ground substance layer (gs) for a very short distance (6 nm). The gs was smooth, up to 500 nm thick, without tubules, and contained a few vesicles. Longitudinally cut bradyzoites at 54 days PI were banana-shaped, 7.8 × 2.2 µm (n = 5). Molecular characterization using 18S rRNA, 28S rRNA, ITS-1, and cox1 genes indicated a close relationship with other Sarcocystis parasites that have snake-rodent life cycles. The parasite in the present study was molecularly and biologically similar to a previously reported isolate (designated earlier as Sarcocystis sp. ex Pantherophis alleghaniensis) from P. alleghaniensis, and it was structurally different from other Sarcocystis species so far described.

A new species of Caryospora Léger, 1904 (Apicomplexa: Eimeriidae) from the Guanabara spotted night snake Siphlophis pulcher (Raddi) (Reptilia: Dipsadidae) in the Atlantic Forest, Brazil.

A new coccidian species of the genus Caryospora Léger, 1904 (Apicomplexa, Eimeriidae) is described based on material from the Guanabara spotted night snake Siphlophis pulcher (Raddi) (Reptilia: Dipsadidae) in a coastal area of the Atlantic Forest in Ilha Grande Island, state of Rio de Janeiro, southeastern Brazil. Caryospora ceadsensis n. sp. possesses spheroidal to slightly subspheroidal oöcysts measuring 17-24 × 17-24 (22.1 × 22.0) µm, with a c.1.3 µm thick bi-layered wall (inner layer smooth, outer layer slightly striated), length/width (L/W) ratio of 1.0-1.1 (1.0) and a highly refractive polar granule. Sporocysts are ellipsoidal, 11-17 × 10-13 (14.7 × 11.2) µm, with a L/W ratio of 1.0-1.5 (1.3), with a nipple-like Stieda body (1.1 µm high and 2.9 µm wide) and a large, bubble-shaped sub-Stieda body 1.7 µm high and 3.8 µm wide (1.0-2.0 × 3.0-4.5 µm). Sporocyst residuum composed of granules of differing sizes. Sporozoites with striations. This is the sixth record of a species of Caryospora in snakes of the family Dipsadidae in Brazil.

A new species of Hepatozoon Miller, 1908 (Apicomplexa: Adelerina) from the snake Philodryas nattereri Steindachner (Squamata: Dipsadidae) in northeastern Brazil.

Based on both unique morphological characteristics of the gamont, distinct changes caused to the host erythrocyte and analysis of partial 18S rRNA gene sequences, a new parasite of the genus Hepatozoon Miller, 1908 is described from the snake Philodryas nattereri Steindachner (Squamata: Dipsadidae) in northeastern Brazil. The new species, Hepatozoon musa n. sp., is characterized by large and curved mature gamonts (18.9 ± 0.9 µm in length and 3.8 ± 0.3 µm in width) that considerably engorge infected host erythrocytes and displace the nucleus laterally, which become longer and thinner. Phylogenetic estimates indicate the new species is more closely related to the recently described Hepatozoon cuestensis O'Dwyer, Moço, Paduan, Spenassatto, Silva & Ribolla, 2013, from Brazilian rattlesnakes. These recent findings highlight the need for further studies of Hepatozoon to better determine the biodiversity of this common but poorly-studied parasite group.

Vertical Transmission of Hepatozoon in the Garter Snake Thamnophis elegans.

Vertical transmission of blood parasites has been demonstrated in humans and some domestic species, but it has not been well documented in wild populations. We assessed whether Hepatozoon blood parasites are vertically transmitted in naturally infected individuals of the viviparous western terrestrial garter snake ( Thamnophis elegans ). Blood smears were taken from nine wild-caught gravid female snakes at capture, preparturition, and postparturition, and then from their laboratory-born offspring at age 2 mo and 1 yr. All infected offspring were born to four infected females, although not all offspring in a given litter were necessarily infected. Parasites were not detected in offspring born to the five uninfected mothers. The highest parasite loads were found in neonates at 2 mo of age. Parasite prevalence did not vary between sexes in offspring, but females showed higher loads than did males when 2 mo old. This study supports vertical transmission of Hepatozoon in naturally infected viviparous snakes and suggests that vertical transmission of hematozoan parasites might be an overlooked mode of transmission in wildlife.

First report of parasitism by Hexametra boddaertii (Nematoda: Ascaridae) in Oxyrhopus guibei (Serpentes: Colubridae).

The current study summarizes the postmortem examination of a specimen of Oxyrhopus guibei (Serpentes, Colubridae) collected in Iguazu National Park (Argentina), and found deceased a week following arrival to the serpentarium of the National Institute of Tropical Medicine (Argentina). Although the snake appeared to be in good health, a necropsy performed following its death identified the presence of a large number of roundworms in the coelomic cavity, with indications of peritonitis and serosal adherence. Additional observations from the necropsy revealed small calcifications in the mesothelium of the coelomic cavity; solid and expressive content in the gallbladder; massive gastrointestinal obstruction due to nematodes; and lung edema and congestion. Histopathological analyses of lung sections also showed proliferative heterophilic and histiocytic pneumonia. Parasites isolated from both the intestine and coelomic cavity were identified as Hexametra boddaertii by a combination of light and scanning electron microscopic examination. Results from this necropsy identify O. guibei as a new host for H. boddaertii, and is the first report of a natural infection by Hexametra in Argentina. Since Hexametra parasites may contribute to several pathological conditions in humans, and with the recent availability of O. guibei specimens through the illegal pet trade, it is necessary to consider the possibility of zoonotic helminth transmission of Hexametra from snake to human.

Ancient, globally distributed lineage of Sarcocystis from sporocysts of the Eastern rat snake (Pantherophis alleghaniensis) and its relation to neurological sequalae in intermediate hosts.

There is an emerging concern that snakes are definitive hosts of certain species of Sarcocystis that cause muscular sarcocystosis in human and non-human primates. Other species of Sarcocystis are known to cycle among snakes and rodents, but have been poorly characterized in the USA and elsewhere. Although neurological sequalae are known for certain species of Sarcocystis, no such neurological symptoms are known to typify parasites that naturally cycle in rodents. Here, sporocysts of a species of Sarcocystis were found in the intestinal contents of a rat snake (Pantherophis alleghaniensis) from Maryland, USA. The sporocysts were orally infective for interferon gamma gene knockout (KO) mice, but not to Swiss Webster outbred mice. The KO mice developed neurological signs, and were necropsied between 33 and 52 days post-inoculation. Only schizonts/merozoites were found, and they were confined to the brain. The predominant lesion was meningoencephalitis characterized by perivascular cuffs, granulomas, and necrosis of the neuropil. The schizonts and merozoites were located in neuropil, and apparently extravascular. Brain homogenates from infected KO mice were infective to KO mice and CV-1 cell line. DNA extracted from the infected mouse brain, and infected cell cultures revealed the highest identity with Sarcocystis species that employ snakes as definitive hosts. This is the first report of Sarcocystis infection in the endangered rat snake (P. alleghaniensis) and the first report of neurological sarcocystosis in mice induced by feeding sporocysts from a snake. These data underscore the likelihood that parasites in this genus that employ snakes as their definitive hosts constitute an ancient, globally distributed monophyletic group. These data also raise the possibility that neurological sequalae may be more common in intermediate hosts of Sarcocystis spp. than has previously been appreciated.

Genetic Identification of Spirometra decipiens Plerocercoids in Terrestrial Snakes from Korea and China.

Human sparganosis is a zoonotic disease caused by infection with larval forms (procercoid/plerocercoid) of Spirometra spp. The purpose of this study was to identify Spirometra spp. of infected snakes using a multiplex PCR assay and phylogenetic analysis of mitochondrial DNA sequence data from the spargana of terrestrial snakes obtained from Korea and China. A total of 283 snakes were obtained that included 4 species of Colubridae comprising Rhabdophis tigrinus tigrinus (n=150), Dinodon rufozonatum rufozonatum (n=64), Elaphe davidi (n=2), and Elaphe schrenkii (n=7), and 1 species of Viperidae, Agkistrodon saxatilis (n=60). The snakes were collected from the provinces of Chungbuk, Chungnam, and Gyeongbuk in Korea (n=161), and from China (n=122). The overall infection rate with spargana was 83% (235/283). The highest was recorded for D. rufozonatum rufozonatum (100%), followed by A. saxatilis (85%) and R. tigrinus tigrinus (80%), with a negative result for E. davidi (0%) and E. schrenkii (0%). The sequence identities between the spargana from snakes (n=50) and Spirometra erinaceieuropaei (KJ599680) or S. decipiens (KJ599679) control specimens were 90.8% and 99.2%, respectively. Pairwise genetic distances between spargana (n=50) and S. decipiens ranged from 0.0080 to 0.0107, while those between spargana and S. erinaceieuropaei ranged from 0.1070 to 0.1096. In this study, all of the 904 spargana analyzed were identified as S. decipiens either by a multiplex PCR assay (n=854) or mitochondrial cox1 sequence analysis (n=50).