Ticks collected from wild and domestic animals and natural habitats in the Republic of Korea.

Ticks were collected from 35 animals from 5 provinces and 3 metropolitan cities during 2012. Ticks also were collected by tick drag from 4 sites in Gyeonggi-do (2) and Jeollabuk-do (2) Provinces. A total of 612 ticks belonging to 6 species and 3 genera were collected from mammals and a bird (n=573) and by tick drag (n=39). Haemaphyalis longicornis (n=434) was the most commonly collected tick, followed by H. flava (158), Ixodes nipponensis (11), Amblyomma testudinarium (7), H. japonica (1), and H. formosensis (1). H. longicornis and H. flava were collected from all animal hosts examined. For animal hosts (n>1), the highest Tick Index (TI) was observed for domestic dogs (29.6), followed by Siberian roe deer (17.4), water deer (14.4), and raccoon dogs (1.3). A total of 402 H. longicornis (adults 86, 21.4%; nymphs 160, 39.8%; larvae 156, 38.9%) were collected from wild and domestic animals. A total of 158 H. flava (n=158) were collected from wild and domestic animals and 1 ring-necked pheasant, with a higher proportion of adults (103, 65.2%), while nymphs and larvae only accounted for 12.7% (20) and 22.2% (35), respectively. Only 7 A. testudinarium were collected from the wild boar (6 adults) and Eurasian badger (1 nymph), while only 5 I. nipponensis were collected from the water deer (4 adults) and a raccoon dog (1 adult). One adult female H. formosensis was first collected from vegetation by tick drag from Mara Island, Seogwipo-si, Jeju-do Province.

Molecular Identification of Haemadipsa rjukjuana (Hirudiniformes: Haemadipsidae) in Gageo Island, Korea.

There are 60 species of blood-feeding land leeches, 50 species belonging to the family Haemadipsidae and 10 species belonging to the family Xerobdellidae. Despite recent papers on the land leeches, their taxonomic identification is not fully understood, especially at a species level. In Korea, there have been no historical records of the terrestrial leeches, but recently an unrecorded blood-feeding land leech was discovered at Gageo-do (Island), Korea. Molecular analysis was used to identify the species of 29 leeches collected from Mt. Dock-Sil in Gageo-do. Conventional PCR was conducted using nuclear 18S rRNA and mitochondrial cytochrome c oxidase subunit 1 (CO1) genetic marker. The 18S rRNA sequences revealed that the leeches share 99.9% identity with Haemadipsa rjukjuana (inhabiting Taiwan), and the CO1 sequences revealed that the leeches are very close to H. rjukjuana (inhabiting Taiwan). The CO1 sequences were separated into 2 categories, 1 with 94.6% and the other with 94.3% similarity to the H. rjukjuana L00115A (inhabiting Taiwan). This new finding of the land leech is the first record in Korea. In addition, the north range of the distribution of the blood-feeding leech (Hirudiniformes: Haemadipisidae) should be reconsidered including Korea.

Climate change induced habitat expansion of nutria (Myocastor coypus) in South Korea.

The nutria, (Myocastor coypus), is a semiaquatic rodent native to the subtropical and temperate regions of South America. The species was introduced to South Korea for meat and fur production purposes and a wild population has become established. The species subsequently invaded aquatic ecosystems and destroyed aquatic vegetation and cultivated crops. Thus, it is essential to understand their current distribution and future range expansion for effective control and eradication strategies to reduce the risk of colonization into new regions. In this study, we used niche modeling procedure to identify potentially suitable habitats for M. coypus under current and future predicted climate change using the maximum entropy algorithm. We found that the main habitat area of M. coypus is expected to expand under a warming climate from ~ 4069 km2 in the southern and southeastern regions of South Korea, to the northern border of the country, with estimated ranges of 21,744 km2, 55,859 km2, and 64,937 km2 by 2030, 2050, and 2070, respectively. The findings of the present study assist in identifying the future distribution and potential dispersion routes of M. coypus in South Korea, which is important for informing the government regarding essential management actions plans at regional and local scales.

Species and sex identification of the Korean goral (Nemorhaedus caudatus) by molecular analysis of non-invasive samples.

Korean long-tailed goral (Nemorhaedus caudatus) is one of the most endangered species in South Korea. However, detailed species distribution and sex ratio data on the elusive goral are still lacking due to difficulty of identification of the species and sex in the field. The primary aim of this study was to develop an economical PCR-RFLP method to identify species using invasive or non-invasive samples from five Korean ungulates: goral (N. caudatus), roe deer (Capreolus pygargus), feral goat (Capra hircus), water deer (Hydropotes inermis) and musk deer (Moschus moschiferus). The secondary aim was to find more efficient molecular sexing techniques that may be applied to invasive or non-invasive samples of ungulate species. We successfully utilized PCR-RFLP of partial mitochondrial cytochrome b gene (376 bp) for species identification, and sex-specific amplification of ZFX/Y and AMELX/Y genes for sexing. Three species (goral, goat and water deer) showed distinctive band patterns by using three restriction enzymes (XbaI, StuI or SspI). Three different sexing primer sets (LGL331/335 for ZFX/Y gene; SE47/48 or SE47/53 for AMELX/Y gene) produced sex-specific band patterns in goral, goat and roe deer. Our results suggest that the molecular analyses of non-invasive samples might provide us with potential tools for the further genetic and ecological study of Korean goral and related species.

A preliminary analysis of the diet composition of overwintering Bean geese (Anser fabalis) and greater white-fronted geese (A. albifrons) in Korea using PCR on fecal samples.

Bean geese (Anser fabalis) and Greater white-fronted geese (Anser albifrons) are the dominant wintering waterfowl in South Korea. Although they are commonly observed in estuaries and rice fields during the winter, the diet composition of the geese during the winter has rarely been studied. In this study, we provide the results from preliminary analyses on the diet of these two geese species overwintering in Daebu Island of South Korea. We used a total of 13 fecal samples from Bean geese (n = 4) and Greater white-fronted geese (n = 9), and performed a BLAST search for the sequences obtained from 87 clones (n = 36 for Bean geese and n = 51 for Greater white-fronted geese). The diet of Bean geese consisted of five families of plants: Caryophyllaceae (75.0%), Poaceae (13.9%), Asteraceae (5.5%), Polygonaceae (2.8%) and Cucurbitacea (2.8%). On the other hand, the diet of Greater white-fronted geese consisted of 6 families of plants: Poaceae (74.5%), Caryophyllaceae (9.8%), Solanacea (5.9%), Portulacaceae (3.9%), Lamiaceae (3.9%) and Brassicaceae (2.0%). We found that plants of the rice family (Poaceae) are important in the diet of wintering geese, especially for Greater white-fronted geese. This knowledge can be used to establish conservation strategies of the geese overwintering in South Korea.

Molecular detection of Bartonella spp. in terrestrial leeches (Haemadipsa rjukjuana) feeding on human and animal blood in Gageo-do, Republic of Korea.

BACKGROUND: Leeches can transmit pathogens and are therefore potentially hazardous to human and animal health. However, only a few studies of diseases transmitted by land leeches have been reported. The purpose of the present study was to analyse which pathogens are carried in Haemadipsa rjukjuana, the first recorded sanguivorous land leech in the Republic of Korea (ROK). FINDINGS: A total of 173 H. rjukjuana were collected from Mt. Dock-Sil on Gageo-do Island, ROK during July 2011. Conventional PCR was conducted for analysis of the origin of blood meal, as well as for detection of species of Anaplasma, Bartonella, Borrelia, Ehrlichia, Rickettsia, and Wolbachia in the leech specimens. Bartonella DNA was detected in eight of the specimens studied based on partial ITS sequence analysis. Seven of the DNA samples were closely related to Bartonella grahamii (99.6-100 % similarity), and one sample exhibited a 90.6 % similarity with Bartonella sp. from Taiwan. Sequences of the mitochondrial cytochrome b gene were generated for a total of 35 of the 173 leech internal organ samples. These included sequences of human (n = 10), mouse (n = 8), weasel (n = 6) and bird (n = 11) origin. Of these 35 sequences, 68.5 % were from mammals, including humans, and 31.4 % were from migratory birds that pass through Gageo-do, ROK. CONCLUSIONS: Although the present study does not provide evidence that leeches indeed transmit Bartonella species to hosts directly, to our knowledge this is the first report on Bartonella DNA being detected from leeches. Therefore, further studies are needed to explore the possibility of zoonotic pathogen transmission by land leeches.

Genetic diversity and phylogeography of Siberian roe deer, Caproulus pygargus, in central and peripheral populations.

Current understanding of phylogeographical structure and genetic diversity of Siberian roe deer remains limited mainly due to small sample size and/or low geographical coverage in previous studies. Published data suggest at least two phylogroups: western (Ural Mountains and Western Siberia) and eastern (east from lake Baikal, including the Korean peninsula), but their phylogenetic relationship remains unclear. Combined sequences of cytochrome b (1140 bp) and the mtDNA control region (963 bp) were analyzed from 219 Siberian roe deer from 12 locations in Russia, Mongolia, and South Korea, which cover a large part of its range, to assess genetic diversity and phylogeographical status. Special emphasis was placed on the demographic history and genetic features of central, peripheral, and isolated populations. Results of median-joining network and phylogenetic tree analyses indicate that Siberian roe deer from the Urals to the Pacific Ocean are genetically diverse and that geographical distribution and composition of haplogroups coincide with previously described ranges of the subspecies Capreolus pygargus pygargus and Capreolus pygargus tianschanicus. We found that peripheral populations in the northwestern parts of the species range (Urals), as well as the isolated population from Jeju Island, are genetically distinct from those in the core part of the range, both in terms of genetic diversity and quantitative composition of haplogroups. We also found that northwestern (Urals) and northern (Yakutia) peripheral populations share the same haplogroup and fall into the same phylogenetic clade with the isolated population from Jeju Island. This finding sheds light on the taxonomic status of the Jeju Island population and leads to hypotheses about the discordance of morphological and genetic evolution in isolated populations and specific genetic features of peripheral populations.

Mitochondrial genetic diversity, phylogeny and population structure of Hydropotes inermis in South Korea.

The water deer (Hydropotes inermis) is one of the rarest species of deer in the family Cervidae. Only two subspecies exist in East Asia, and few studies have examined the genetic characteristics of the species. Here, we investigated the genetic diversity, phylogeny and population differentiation of the Korean subspecies (H. inermis argyropus). Seventeen mitochondrial D-loop haplotypes (822 bp) were detected and analyzed from 107 individual samples, together with a Chinese subspecies (H. inermis inermis) haplotype. The genetic diversity of the Korean subspecies is lower (π = 0.756%, h = 0.867) than that of the Chinese subspecies estimated in a previous study. This low genetic diversity may result from historical anthropogenic disturbances and/or a founder effect during the glacial period. The phylogenetic tree and median-joining network showed no location-specific distribution of D-loop haplotypes, but revealed two major lineages, A and B, of water deer. The A and B lineages were separated from each other at the beginning of the Pleistocene era (2.1-1.3 million years ago), with a genetic divergence of 1.332 ± 0.340%. The genetic divergence within lineages A and B was 0.525 ± 0.167% and 0.264 ± 0.113%, respectively. This suggests that climate change affected the division of the two lineages. Water deer sampled from the three Korean regions (26 locations) were slightly distinct in their genetic structure (AMOVA: F(ST) = 0.28416, P < 0.00001; Φ(ST) = 0.19239, P < 0.00001). Such slight population differentiation may be derived from differential dispersal ability in males and females. The use of genetic markers, such as nuclear microsatellite and Y-linked DNA markers, and samples collected from various localities in East Asia should improve our understanding of the water deer's genetic characteristics.

First report for the seasonal and annual prevalence of flea-borne bartonella from rodents and soricomorphs in the Republic of Korea.

Rodents and soricomorphs are animal hosts of fleas and associated zoonotic microbial pathogens. A total of 4,889 small mammals were collected from Gyeonggi and Gangwon Provinces, Republic of Korea, from 2008 through 2010, including: Apodemus agrarius (4,122, 84.3%), followed by Crocidura lasiura (282, 5.8%), Microtus fortis (257, 5.3%), Myodes regulus (77, 1.6%), Micromys minutus (71, 1.5%), Mus musculus (63, 1.3%), and 4 other species (17, 0.3%). A total of 1,099 fleas belonging to 10 species and 7 genera were collected. Ctenophthalmus congeneroides (724, 65.9%) was the most commonly collected flea, followed by Stenoponia sidimi (301, 27.4%), Neopsylla bidentatiformis (29, 2.6%), and Rhadinopsylla insolita (25, 2.3%). The remaining species accounted for only 1.8% (20, range 1-6) of all fleas collected. The 2 dominant flea species, C. congeneroides and S. sidimi, showed an inverse seasonal pattern, with higher populations of C. congeneroides from January-September, whereas S. sidimi was more frequently collected during October-December. The overall flea infestation rates (FIR) and flea indices (FI) were 14.1% and 0.22, respectively, and were highest during April-June (19.7% and 0.30, respectively). A total of 735 of the 1,099 fleas were assayed for the detection of Bartonella spp. by PCR using Bartonella-specific primers, of which 515 were positive for Bartonella, with an overall maximum likelihood estimate (MLE) of 700.7/1,000. The highest MLE values were observed during April-June (899.2) and July-September (936.2) trapping periods and, although lower, were similar for January-March (566.7) and October-December (574.1). C. congeneroides demonstrated high MLEs for all seasons (range 752.5-934.8), while S. sidimi was positive for Bartonella only during January-March (MLE=342.1) and October-December (MLE=497.2) collection periods. Continued long-term surveillance of small mammals and associated ectoparasites is needed to improve our understanding of the prevalence of Bartonella spp. in fleas and the role of fleas in the zoonotic maintenance and transmission of Bartonella to humans.