Morphologic and Genetic Identification of Diphyllobothrium nihonkaiense in Korea

Diphyllobothrium nihonkaiense was first described by Yamane in 1986 but the taxonomical features have been obscure due to lack of critical morphologic criteria in its larval and adult stages. In Korea, this tapeworm had long been known as Diphyllobothrium latum. In this study, we observed 62 specimens collected from Korean residents and analyzed them by morphological features and nucleotide sequences of mitochondrial cox1 gene as well as the ITS1 region. Adult tapeworms were examined after carmine or trichrome stain. Longitudinal sections of the gravid proglottids showed an obtuse angle of about 150 degree between the cirrus sac and seminal vesicle. This angle is known as a major differential point compared with that of D. latum. Nucleotide sequence differences between D. latum and the specimens from Koreans represented 17.3% in mitochondrial DNA cox1 gene. Sequence divergence of ITS1 among 4 Korean isolates was 0.3% and similarity was 99.7% with D. nihonkaiense and D. klebanovskii. All of the Korean specimens analyzed in this study were identified as being D. nihonkaiense (n = 62). We propose its Korean name as "Dong-hae-gin-chon-chung" which means ''long tapeworm of the East Sea'' for this newly analyzed diphyllobothriid tapeworm in Korea.     

<Emphasis Type="Italic">Corynosoma australe</Emphasis> Johnston, 1937 and <Emphasis Type="Italic">C. cetaceum</Emphasis> Johnston &; Best, 1942 (Acanthocephala: Polymorphidae) from marine mammals and fishes in Argentinian waters: allozyme markers and taxonomic status

Genetic and morphological studies were carried out on acanthocephalans belonging to Corynosoma Lühe, 1904 and referable to the species C. cetaceum Johnston & Best, 1942 and C. australe Johnston, 1937, which were recovered from both definitive and intermediate hosts in Argentinian waters. The aims were to estimate the level of genetic differentiation between the two taxa at any stage of their life-cycle, to provide genetic (allozyme) markers for their recognition and to analyse the systematic status of both taxa. Acanthocephalans were collected from the stomach and intestine of Arctocephalus australis (Zimmerman), the intestine of Mirounga leonina (Linnaeus) and the stomach of Pontoporia blainvillei Gervais & D’Orbigny (definitive hosts) in Argentinian waters. Alternative alleles at all the 13 enzymatic loci studied were observed for C. australe and C. cetaceum. The specimens from the stomach of both P. blainvillei and A. australis were identified, on the basis of the great number of diagnostic loci found, as C. cetaceum; those from intestine of both A. australis and M. leonina as C. australe. A high level of genetic differentiation (D_Nei=∞: I_Nei = 0.00) between the two taxa was found, suggesting a generic distinction between the two species. Cystacanths of the two species from the body-cavity of the fish Cynoscion guatucupa (Cuvier) collected from the same geographical area were identified genetically. Morphological patterns, such as the number of hooks and hook rows on the proboscis, the distribution of somatic and genital armature, and other morphometric and meristic differences, in addition to ecological data, enabled the identification of these two species at cystacanth, juvenile and adult stages. However, a number of morphological and morphometric features of the Argentinian material were different to those of C. australe and C. cetaceum described from other regions of the world.     

DNA barcoding and phylogenetic analysis of midges belonging to Culicoides (Diptera: Ceratopogonidae) subgenus Hoffmania in Yunnan,China

DNA barcodes obtained from cytochrome c oxidase subunit 1 (cox1) offer a fast and easy way to identify a range of biological organisms. Culicoides (Diptera: Ceratopogonidae) are a group of small, blood sucking midges whose species are the vectors for some arboviruses, such as bluetongue virus, African horse sickness virus, epizootic hemorrhagic disease virus and equine encephalosis virus. Identification of these small insects is difficult so constructing DNA barcode libraries for species present in certain areas is helpful to clarify the taxonomy and assist non-specialist workers to identify species. In this study, we analysed specimens belonging to C. subgenus Hoffmania collected from 12 towns of Yunnan Province, China. Specimens were identified by morphology and processed to construct DNA barcodes. A total of 185 specimens referable to 6 morphological species were processed for cox1 and 28S rRNA sequencing. The resulting 185 cox1 sequences were assigned to 13 barcode index numbers (BINs) which include 9 novel BINs. Molecular and morphological evidence was used to support the transfer of 4 species previously assigned to C. subg. Avaritia into C. subg. Hoffmania. Molecular analysis revealed the presence of 7 potential cryptic species within C. innoxius, three within C. liui and two within C. insignipennis.     

Molecular and morphological evidence suggests the reallocation from Parastrigea brasiliana (Szidat, 1928) Dubois, 1964 to Apharyngostrigea Ciurea, 1927 (Digenea: Strigeidae), a parasite of boat-billed heron (Cochlearius cochlearius) from the Neotropical region

Parastrigea brasiliana (Szidat, 1928) Dubois, 1964, was described from (Cochlearius cochlearius) in South America. The taxonomy of this species has been unstable due that it was described as a member of Strigea Abildgaard, 1790. However, the same author one year later transferred it to Apharyngostrigea Ciurea, 1927 and since then, it has been alternatively placed in the genus Apharyngostrigea or Parastrigea Szidat, 1928 from Strigeidae. In the current research, specimens identified as P. brasiliana were collected from type host in southeastern Mexico. We sequenced three molecular markers: the internal transcribed spacers ITS1 and ITS2 including the 5.8S gene (ITS region), the D1-D3 domains of the large subunit (LSU) from nuclear DNA and cytochrome c oxidase subunit I (cox 1) from mitochondrial DNA. These sequences were aligned with other sequences available in the GenBank dataset from Strigeidae. Maximum likelihood and Bayesian analyses inferred with three molecular markers consistently showed that P. brasiliana is not closely related to other members of the genus Parastrigea and are placed in a reciprocal monophyletic clade inside Apharyngostrigea, with very low genetic divergence, varying from 0 to 0.09% for the ITS, from 0 to 0.08% for the LSU and from 0.21 to 0.43% for cox 1. Consequently, we proposed to reallocate it to A. brasiliana. The phylogenetic analyses obtained are key and very useful for re-evaluate the morphology of A. brasiliana because this species share morphological characters with the genera Parastrigea (concentration of vitelline follicles distributed in two lateral expansions on the forebody) and Apharyngostrigea (absence of pharynx). Finally, the current record of A. brasiliana expands its distribution range in four countries, namely, the USA, Mexico, Venezuela and Brazil, in the Neotropical region.     

Cat fleas (Ctenocephalides felis) carrying Rickettsia felis and Bartonella species in Hong Kong

Fleas are commonly recorded on stray as well as domestic dogs and cats in Hong Kong. Fleas can be a major cause of pruritus in dogs and cats and also vectors of potentially zoonotic bacteria in the genera Rickettsia and Bartonella. Morphological examination of 174 fleas from dogs and cats living in Hong Kong revealed only cat fleas (Ctenocephalides felis). Cytochrome c oxidase subunit 1 gene (cox1) genotyping of 20 randomly selected specimens, revealed three cox1 haplotypes (HK-h1 to HK-h3). The most common haplotype was HK-h1 with 17 specimens (17/20, 85%). HK-h1 was identical to cox1 sequences of fleas in Thailand and Fiji. HK-h1 and HK-h2 form a distinct cat flea cox1 clade previously recognized as the Clade 3. HK-h3 forms a new Clade 6. A multiplex Bartonella and Rickettsia real-time PCR of DNA from 20 C. felis found Bartonella and Rickettsia DNA in three (15%) and ten (50%) C. felis, respectively. DNA sequencing confirmed the presence of R. felis, B. clarridgeiae and Bartonella henselae. This is the first reported study of that kind in Hong Kong, and further work is required to expand the survey of companion animals in the geographical region. The sampling of fleas on domestic cats and dogs in Hong Kong revealed them to be exclusively infested by the cat flea and to be harbouring pathogens of zoonotic potential.     

PHYLOGEOGRAPHY OF CUTLERIA CYLINDRICA (CUTLERIALES,PHAEOPHYCEAE) IN NORTHEASTERN ASIA,AND THE IDENTITY OF AN INTRODUCED POPULATION IN CALIFORNIA1

Cutleria cylindrica Okamura was described from Japan in 1902 and has been reported only from northwestern Asia until its relatively recent discovery in California, USA, and Baja California, Mexico. To clarify the genetic relationships within and among the disjunct populations, we carried out a molecular phylogenetic study, as well as the examination of sex ratio and the life‐history patterns, of populations in Japan, Korea, and California. Based on the DNA sequences of mitochondrial genes cox2, cox3, the open reading frame (ORF) region, and the spacer between cox3 and ORF, a total of 23 haplotypes were detected in the 85 individuals from 20 localities in Japan, Korea, and California. All localities in Japan and Korea included multiple haplotypes, but only a single haplotype was found in California. There was a positive relationship between distance and genetic divergence in Japan and Korea. The single haplotype found in California was the same as one occurring in Japan (Aomori Pref. and Fukuoka Pref.) and Korea (Daedaepo, Pusan). Both male and female gametophytes were distributed in most northeastern Asian populations. Only female gametophytes, developing parthenogenetically from female gametes, were found in California and Aomori Pref., Japan. On the basis of these results, we conclude that the disjunct population of C. cylindrica in California originated from a relatively recent introduction from Japan and shares its origin with the parthenogenetic population in the Tsugaru Strait.     

A Revision of the genus Wislizenia (Capparidaceae) based on population studies

Morphological and cytological studies of mass collections from the southwestern United States and Baja California, and morphological studies on herbarium specimens from these areas and northern Mexico have been used to solve a taxonomic problem within the genusWislizenia. Canonical analysis of morphological data from mass collections separates three geographic groups from: California, Baja California, and states east of California. A similar analysis of herbarium specimens collected throughout the range ofWislizenia refracta confirms the reality of these groups, for which the status of subspecies is proposed: 1) subsp.californica from the California central valley, 2) subsp.palmeri from Baja California and the Sonoran coast, and 3 ) subsp.refracta from states east of California. This taxonomic treatment differs from any of the varied previous ones. Chromosome numbers for six of the mass collections were found to ben = 20, in agreement with other reports.     

88 Preliminary observations on the identity of the red algal genus Caloglossa (delesseriaceae,ceramiales) in the northwestern gulf of mexico

Phylogenetic analyses inferred from rbcL sequences indicates that specimens of Caloglossa (Harvey) G. Martens collected from Vermilion Bay, Louisiana and Galveston, Texas, reported as C. leprieurii (Montagne) G. Martens (1869) constitutes a separate taxon. Studies are presently being conducted to compare this northwestern Gulf of Mexico taxon with material of C. intermedia Kamiya et West in Kamiya et al. (2000, J. Phycol. 36:411‐420) from the Western Atlantic coast and with C. apomeiotica West et Zuccarello in West et al. (1994, Bot. Mar. 37:381–390) from Baja California, Mexico.     

Sympatric Distribution of Three Human Taenia Tapeworms Collected between 1935 and 2005 in Korea

Taeniasis has been known as one of the prevalent parasitic infections in Korea. Until recently, Taenia saginata had long been considered a dominant, and widely distributed species but epidemiological profiles of human Taenia species in Korea still remain unclear. In order to better understand distribution patterns of human Taenia tapeworms in Korea, partial nucleotide sequences of mitochondrial cox1 and ITS2 (internal transcribed spacer 2) were determined, along with morphological examinations, on 68 Taenia specimens obtained from university museum collections deposited since 1935. Genomic DNA was extracted from formalin-preserved specimens. Phylogenetic relationships among the genotypes (cox1 haplotype) detected in this study were inferred using the neighbor-joining method as a tree building method. Morphological and genetic analyses identified 3 specimens as T. solium, 51 specimens as T. asiatica, and 14 specimens as T. saginata. Our results indicate that all 3 Taenia tapeworms are sympatrically distributed in Korea with T. asiatica dominating over T. saginata and T. solium.     

Molecular Characterization of Taenia multiceps Isolates from Gansu Province,China by Sequencing of Mitochondrial Cytochrome C Oxidase Subunit 1

A total of 16 Taenia multiceps isolates collected from naturally infected sheep or goats in Gansu Province, China were characterized by sequences of mitochondrial cytochrome c oxidase subunit 1 (cox1) gene. The complete cox1 gene was amplified for individual T. multiceps isolates by PCR, ligated to pMD18T vector, and sequenced. Sequence analysis indicated that out of 16 T. multiceps isolates 10 unique cox1 gene sequences of 1,623 bp were obtained with sequence variation of 0.12-0.68%. The results showed that the cox1 gene sequences were highly conserved among the examined T. multiceps isolates. However, they were quite different from those of the other Taenia species. Phylogenetic analysis based on complete cox1 gene sequences revealed that T. multiceps isolates were composed of 3 genotypes and distinguished from the other Taenia species.     

Phylogeny of Veronica in the Southern and Northern Hemispheres based on plastid,nuclear ribosomal and nuclear low-copy DNA

The cosmopolitan and ecologically diverse genus Veronica with approximately 450 species is the largest genus of the newly circumscribed Plantaginaceae. Previous analyses of Veronica DNA sequences were in stark contrast to traditional systematics. However, analyses did not allow many inferences regarding the relationship between major groups identified, hindering further analysis of diversification and evolutionary trends in the genus. To resolve the backbone relationships of Veronica, we added sequences from additional plastid DNA regions to existing data and analyzed matching data sets for 78 taxa and more than 5000 aligned characters from nuclear ribosomal DNA and plastid DNA regions. The results provide the best resolved and supported estimate of relationships among major groups in the Northern (Veronica s. str.) and Southern Hemisphere (hebes). We present new informal names for the five main species groups within the Southern Hemisphere sect. Hebe. Furthermore, in two instances we provide morphological and karyological characters supporting these relationships. Finally, we present the first evidence from nuclear low-copy CYCLOIDEA2-region to compare results from the plastid genome with the nuclear genome.     

Isolation of the pyrrolizidine alkaloid 1-epialexine from Castanospermum australe

The Australian leguminous tree Castanospermum australe contains the anti-viral glucose analogue indolizidine alkaloid castanospermine. As the result of a search for new bioactive carbohydrate-like compounds we now report the isolation of the novel polyhydroxylated pyrrolizidine alkaloid, 1-epialexine, from the leaves and stems of C. australe. 1-Epialexine is a weak inhibitor of β-mannosidase from Cellullomonas fimi.     

Genotypic characterization and species identification of Fasciola spp. with implications regarding the isolates infecting goats in Vietnam

Ribosomal RNA sequences (361 or 362 bp) of the second internal transcribed spacer 2 (ITS-2) and a portion of mitochondrial cox1 (423 bp) for Fasciola spp. obtained from specimens collected in indigenous and hybrid goats and sheep in Vietnam were characterized for genotypic status and hybridization/introgression. Alignment of 48 ITS-2 sequences (also those from goats and sheep in this study) indicates that F. gigantica and F. hepatica differ typically from each other at seven sites whereas one of these is a distinguishing deletion (T) at the 327th position in F. gigantica relative to F. hepatica. The isolates from the mountainous goats in the North of Vietnam (Yen Bai province) showed the ITS-2 composition relatively identical to that of F. hepatica. The ITS-2 sequences from populations of Fasciola isolates in goats had probably experienced introgression/hybridization as reported previously in other ruminants and humans. All Vietnamese goat-of-origin specimens had high pairwise percentage of mitochondrial cox1 sequences to F. gigantica (97-100%), and very low identity to F. hepatica (91-93%), suggesting their maternal linkage to be traced to F. gigantica. The presence of hybrid and/or introgressed populations of liver flukes bearing genetic material from both F. hepatica and F. gigantica in the goats/sheep in Vietnam, regardless of indigenous or imported hosts, appears to be the first demonstration from a tropical country.     

Monitoring of Fasciola Species Contamination in Water Dropwort by cox1 Mitochondrial and ITS-2 rDNA Sequencing Analysis

Fascioliasis, a food-borne trematode zoonosis, is a disease primarily in cattle and sheep and occasionally in humans. Water dropwort (Oenanthe javanica), an aquatic perennial herb, is a common second intermediate host of Fasciola, and the fresh stems and leaves are widely used as a seasoning in the Korean diet. However, no information regarding Fasciola species contamination in water dropwort is available. Here, we collected 500 samples of water dropwort in 3 areas in Korea during February and March 2015, and the water dropwort contamination of Fasciola species was monitored by DNA sequencing analysis of the Fasciola hepatica and Fasciola gigantica specific mitochondrial cytochrome c oxidase subunit 1 (cox1) and nuclear ribosomal internal transcribed spacer 2 (ITS-2). Among the 500 samples assessed, the presence of F. hepatica cox1 and 1TS-2 markers were detected in 2 samples, and F. hepatica contamination was confirmed by sequencing analysis. The nucleotide sequences of cox1 PCR products from the 2 F. hepatica-contaminated samples were 96.5% identical to the F. hepatica cox1 sequences in GenBank, whereas F. gigantica cox1 sequences were 46.8% similar with the sequence detected from the cox1 positive samples. However, F. gigantica cox1 and ITS-2 markers were not detected by PCR in the 500 samples of water dropwort. Collectively, in this survey of the water dropwort contamination with Fasciola species, very low prevalence of F. hepatica contamination was detected in the samples.     

A re-evaluation of morphological characters of the invasive ascidian Corella eumyota reveals two different species at the tip of South America and in the South Shetland Islands, Antarctica

The native solitary ascidian Corella eumyota Traustedt, 1882) is commonly found in the Southern Hemisphere in the cold-temperate waters of the Subantarctic and Antarctic regions. Its recent spread into the Northern Hemisphere throughout the NE Atlantic gave the species the status of invasive. Together with its widespread distribution, reports on its wide variability (several distinct morphological characters, genetic discontinuities and also possible misidentifications) cast doubt on the taxonomic status of different populations of this species. This work, based on the observation, quantification and analysis of specific morphological characters in specimens collected at five different localities of South America and Antarctica, strongly indicates that there are two different species: C. eumyota from South America and Corella antarctica Sluiter, 1905) from Antarctica, which has been till now considered a junior synonym of the former. The species clearly differ in the arrangement of the gonadal ducts, the size of the larvae and the shape of the anus, among other characters. Morphological variation displays a defined, discrete grouping supporting a clear differentiation into two species. This result shows the need for careful inspection of specimens to avoid wrong interpretations in a context of changes of marine biota due to biological invasions.     

The genus Platychara from the Western Hemisphere

The systematics of four species belonging to the genus Platychara (Charophyta) from the Western Hemisphere is discussed. Three of the species, as defined herein, occur in Cretaceous and Paleocene rocks from Mexico through South America. The type species, P. compressa (Peck and Reker) Grambast, also of Cretaceous and Paleocene age, is herein restricted to deposits north of Mexico. These latter restrictions geographically separate P. compressa and P. perlata as presently defined but the relationship between these two species is still uncertain. A new species, P. grambastii, is proposed for specimens from Maestrichtian sediments in Jamaica.     

Development of microsatellite markers for the white mangrove (Laguncularia racemosa C.F. Gaertn., Combretaceae)

We developed nine new microsatellite markers for the white mangrove (Laguncularia racemosa C.F. Gaertn.), a mangrove species widely distributed in the Western Hemisphere. The loci were developed from a sample collected in Baja California Sur, Mexico, and were screened for polymorphism in 108 individuals from the Pacific Coast of Mexico and Costa Rica. The level of polymorphism was relatively low, from two to four alleles. These are the first set of polymorphic markers described for L. racemosa and should be of value in studying regional genetic differentiation among natural populations of this species.     

Rediscovery of Polypodium calirhiza (Polypodiaceae) in Mexico

This study addresses reported discrepancies regarding the occurrence of Polypodium calirhiza in Mexico. The original paper describing this taxon cited collections from Mexico, but the species was omitted from the recent Pteridophytes of Mexico. Originally treated as a tetraploid cytotype of P. californicum, P. calirhiza now is hypothesized to have arisen through hybridization between P. glycyrrhiza and P. californicum. The tetraploid can be difficult to distinguish from either of its putative parents, but especially so from P. californicum. Our analyses show that a combination of spore length and abaxial rachis scale morphology consistently distinguishes P. calirhiza from P. californicum, and we confirm that both species occur in Mexico. Although occasionally found growing together in the United States, the two species are strongly allopatric in Mexico: P. californicum is restricted to coastal regions of the Baja California peninsula and neighboring Pacific islands, whereas P. calirhiza grows at high elevations in central and southern Mexico. The occurrence of P. calirhiza in Oaxaca, Mexico, marks the southernmost extent of the P. vulgare complex in the Western Hemisphere.