Integrative taxonomy of Mesocriconema onoense (Tylenchida: Criconematidae) from Vietnam highly suggests the synonymization of Mesocriconema brevistylus and related species.

The genus Mesocriconema is one of the most diverse genera within the family Criconematidae, known as ring nematodes, with more than 90 species. Although species in this genus usually show distinct morphological characterizations, the identification based only on morphology can lead to misidentification in many studies resulted in a number of synonymizations in the genus over time. In this study, an integrated approach has been applied in characterizing Mesocriconema onoense from Vietnam. The molecular data of 28S rRNA, ITS, 18S rRNA regions were analyzed and discussed to confirm the correct names on GenBank. Besides, phylogenetic analyses of 28S rRNA, ITS, and 18S rRNA regions of Mesocriconema species revealed that Mesocriconema brevistylus should be considered as a junior synonym of M. onoense. Consequently, M. helicus, M. onostris, and M. paronostris should also be considered as the synonyms of M. onoense.

Characterization of CRISPR Spacer and Protospacer Sequences in Paenibacillus larvae and Its Bacteriophages.

The bacterium Paenibacillus larvae is the causative agent of American foulbrood, the most devastating bacterial disease of honeybees. Because P. larvae is antibiotic resistant, phages that infect it are currently used as alternative treatments. However, the acquisition by P. larvae of CRISPR spacer sequences from the phages could be an obstacle to treatment efforts. We searched nine complete genomes of P. larvae strains and identified 714 CRISPR spacer sequences, of which 384 are unique. Of the four epidemiologically important P. larvae strains, three of these have fewer than 20 spacers, while one strain has over 150 spacers. Of the 384 unique spacers, 18 are found as protospacers in the genomes of 49 currently sequenced P. larvae phages. One P. larvae strain does not have any protospacers found in phages, while another has eight. Protospacer distribution in the phages is uneven, with two phages having up to four protospacers, while a third of phages have none. Some phages lack protospacers found in closely related phages due to point mutations, indicating a possible escape mechanism. This study serve a point of reference for future studies on the CRISPR-Cas system in P. larvae as well as for comparative studies of other phage-host systems.

Analysis of accessible chromatin landscape in the inner cell mass and trophectoderm of human blastocysts.

Early embryonic development is characterized by drastic changes in chromatin structure that affects the accessibility of the chromatin. In human, the chromosome reorganization and its involvement in the first linage segregation are poorly characterized due to the difficulties in obtaining human embryonic material and limitation on low input technologies. In this study, we aimed to explore the chromatin remodeling pattern in human preimplantation embryos and gain insight into the epigenetic regulation of inner cell mass (ICM) and trophectoderm (TE) differentiation. We optimized ATAC-seq (an assay for transposase-accessible chromatin using sequencing) to analyze the chromatin accessibility landscape for low DNA input. Sixteen preimplantation human blastocysts frozen on Day 6 were used. Our data showed that ATAC peak distributions of the promoter regions (<1 kb) and distal regions versus other regions were significantly different between ICM versus TE samples (P < 0.01). We detected that a higher percentage of accessible binding loci were located within 1 kb of the transcription start site in ICM compared to TE (P < 0.01). However, a higher percentage of accessible regions was detected in the distal region of TE compared to ICM (P < 0.01). In addition, eight differential peaks with a false discovery rate <0.05 between ICM and TE were detected. This is the first study to compare the landscape of the accessible chromatin between ICM and TE of human preimplantation embryos, which unveiled chromatin-level epigenetic regulation of cell lineage specification in early embryo development.

Assessment of the diversity of Brazilian entomopathogenic fungi in the genus Beauveria.

We combined matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS) along with sequencing of the B locus intergenic region (Bloc) to assess the diversity of Brazilian species within the anamorphic genus Beauveria. A total of 121 strains maintained in a government-owned culture collection and isolated from a range of hosts/substrates over a long time span (1981-2015) were assessed. Strains were collected in five of six Brazilian biomes, mostly in the Atlantic Forest (42.2%) and Cerrado (29.8%), primarily from insect pests of crops. All strains were subjected to MS, and those not accurately identified by this technique were genomically analyzed. Among the outcomes of this study, four taxa from the genus Beauveria were recognized, with the great majority of strains belonging to B. bassiana s.str. (93.4%), followed by B. caledonica (2.5%), B. pseudobassiana (2.5%) and B. amorpha (1.6%). B. bassiana s.str. was found in all biomes and isolated from a wide range of hosts/substrates. Due to low numbers, associations of the remaining Beauveria species with specific hosts or habitats/biomes were not clear, except that all three B. caledonica strains were found only in the Cerrado biome and were associated with adults of the banana weevil, Cosmopolites sordidus (Col.:Curculionidae). B. pseudobassiana is reported for the first time on the South American continent, in a subtropical region and from two insect orders not yet associated with this taxon. We also showed that some strains previously ascribed to B. brongniartii were misidentifications. The biodiversity of Beauveria analyzed in our study was comparatively low. The geographic origins of strains used in our study were biased towards biomes with intense human interventions. Future surveys on more conserved, less environmentally disturbed biomes, such as Caatinga, Pampa, Pantanal, and Amazon are needed for a more comprehensive picture of the diversity of Beauveria and related genera in Brazil.

High prevalence of Schistosoma haematobium × Schistosoma bovis hybrids in schoolchildren in Côte d'Ivoire.

Schistosomiasis is a neglected tropical disease, though it is highly prevalent in many parts of sub-Saharan Africa. While Schistosoma haematobium-bovis hybrids have been reported in West Africa, no data about Schistosoma hybrids in humans are available from Côte d'Ivoire. This study aimed to identify and quantify S. haematobium-bovis hybrids among schoolchildren in four localities of Côte d'Ivoire. Urine samples were collected and examined by filtration to detect Schistosoma eggs. Eggs were hatched and 503 miracidia were individually collected and stored on Whatman® FTA cards for molecular analysis. Individual miracidia were molecularly characterized by analysis of mitochondrial cox1 and nuclear internal transcribed spacer 2 (ITS 2) DNA regions. A mitochondrial cox1-based diagnostic polymerase chain reaction was performed on 459 miracidia, with 239 (52.1%) exhibiting the typical band for S. haematobium and 220 (47.9%) the S. bovis band. The cox1 and ITS 2 amplicons were Sanger sequenced from 40 randomly selected miracidia to confirm species and hybrids status. Among the 33 cox1 sequences analysed, we identified 15 S. haematobium sequences (45.5%) belonging to seven haplotypes and 18 S. bovis sequences (54.5%) belonging to 12 haplotypes. Of 40 ITS 2 sequences analysed, 31 (77.5%) were assigned to pure S. haematobium, four (10.0%) to pure S. bovis and five (12.5%) to S. haematobium-bovis hybrids. Our findings suggest that S. haematobium-bovis hybrids are common in Côte d'Ivoire. Hence, intense prospection of domestic and wild animals is warranted to determine whether zoonotic transmission occurs.

Integrative taxonomy of Abacarus mites (Eriophyidae) associated with hybrid sugarcane plants, including description of a new species.

Phytophagous mites belonging to the Eriophyoidea are extremely diverse and highly host-specific. Their accurate morphological identification is hampered by their reduced size and simplified bodies and by the existence of cryptic species complexes. Previous studies have demonstrated the urgency of applying multisource methods to accurate taxonomic identification of eriophyoid mites, especially species belonging to the genus Abacarus. This genus comprises 65 species, of which 37 are associated with grasses and four with sugarcane Saccharum (Poaceae). Recently, Abacarus specimens very similar to Abacarus sacchari were collected from the sugarcane crop in Brazil; however, their taxonomic placement was uncertain. In this study, we used an integrative approach to determine whether A. aff. sacchari specimens belong to A. sacchari or constitute a cryptic species. Morphological data were combined with molecular phylogeny based on the nucleotide sequences of three markers, one mitochondrial (COI) and two nuclear (D2 region of 28S and ITS). Morphological differences were observed between A. aff. sacchari, A. sacchari and A. doctus. The phylogenetic relationships among these three taxa and the genetic distances separating them revealed an interspecific divergence. The results of the morphological and molecular methods were congruent and supported the existence of a new species: Abacarus neosacchari n. sp. Duarte and Navia, herein described. This species belongs to the Abacarus cryptic species complex associated with sugarcane in the Americas. The results of this study, presenting the occurrence of multiple Abacarus species associated with sugarcane, contribute to the knowledge on plants and mites diversity by adding up one more clue highlighting that plant hybridization can be an important mechanism contributing to the speciation of plant-feeding arthropods.

Multi-locus phylogenetic analysis groups the New World bacterium Rickettsia sp. strain ApPR with the Old World species R. africae; proposal of "Candidatus Rickettsia paranaensis".

Rickettsia parkeri sensu stricto (s.s.) is an emerging human pathogen in the Americas. Comprehension of the etiology of R. parkeri infections in South America is complicated by the existence of genetic variants (Atlantic rainforest, NOD and Parvitarsum) of this species that are associated with specific groups of Amblyomma ticks. The rickettsial bacterium strain ApPR was first reported in Amblyomma parkeri ticks in Southern Brazil in 2012 and was considered, based on sequencing of fragments of the gltA, htrA, ompA and ompB genes, to represent yet another genetic variant of R. parkeri. In the current work, a multi-locus phylogenetic analysis employing additional genes and intragenic regions was performed using DNA extracted from (a) larvae of A. parkeri and Amblyomma species haplotype Nazaré ticks collected from wild birds, (b) a nymph of Amblyomma sp. haplotype Nazaré recovered from a monkey (Callicebus nigrifons), representing the first report of that tick parasitizing a non-human primate and (c) from a cultured isolate of ApPR, isolated from colony-reared adults of Amblyomma geayi. Phylogenetic inference performed using Maximum-likelihood (ML), Maximum Parsimony (MP) and Bayesian (B) methods, consistently placed strain ApPR outside the New World R. parkeri complex and instead grouped it in proximity to the Old World species Rickettsia africae and Rickettsia sibirica. Estimates of evolutionary divergence provided additional support for the inferred phylogenetic relationship. Given the clear evolutionary distance between strain ApPR and R. parkeri we propose the recognition of "Candidatus Rickettsia paranaensis".

Salmonella serotype assignment by sequencing analysis of intergenic regions of ribosomal RNA operons.

Salmonella laboratorial detection is usually carried out by bacteriological culture and serological methods. Salmonella isolates are then classified into more than 2,650 serotypes with different somatic (O) and flagellar (H) antigenic combinations. More recently, DNA analysis methods were developed and applied for the identification of Salmonella serotypes, including intergenic spacer regions (ISRs) that separates DNA-encoding ribosomal subunits (rRNA gene) in Salmonella genomes. The present study aimed to evaluate the nucleotide diversity of the ISRs in 2 rRNA operons (rrnB and rrnH) for the assignment of Salmonella serotypes. A total of 63 Salmonella isolates (bacterial cultures) from 21 serotypes were obtained in the period of 2014 to 2017. DNA was extracted, and PCRs were used to detect the genus Salmonella and 4 important serotypes: Enteritidis, Gallinarum, Heidelberg, and Typhimurium. ISRs of the operons rrnB and rrnH were amplified by PCR and then sequenced. All sequence results were submitted to BLASTn search and were aligned in comparison to 72 Salmonella reference nucleotide sequences. The results demonstrated that 60 (95.2%) samples returned a sequence of the same serotype (determined by the traditional serological procedure) after searching in BLASTn and/or in the alignment with the reference sequences for both operons (rrnB and rrnH). These PCR-sequencing procedures had a general agreement index of 0.792 based on the Kappa analysis, 98.7% sensitivity value, 100% specificity, and 98.4% accuracy. Three different phylogenetic trees were generated from the alignments with the sequences of rrnH, rrnB, and rrnH plus rrnB and isolates clustered in specific branches according to the serotypes.

Genetic Characterization of Fungi Isolated from Environmental Samples: A Molecular Surveillance Study of Public Health Importance.

Background: A multistate fungal meningitis outbreak began in September 2012 that affected 751 individuals who received contaminated spinal injections across 20 states in the United States, which led to 64 deaths. In our previous study, we examined 26 environmental swab samples collected from various locations of the manufacturing premises of the compounding company to determine the possible cause of this outbreak and identified 14 novel species of fungi. Objective: In this follow-up study, a total of 198 environmental surveillance samples were collected and analyzed to detect pathogenic fungal species from other compounding company premises located in three regions of the United States. Methods: DNA sequencing was performed at the large subunit ribosomal RNA (LSU rRNA) and internal transcribed spacer (ITS) regions on the 25 positive fungal isolates. Results: Sequence analysis of the ITS1, the ITS2, and the LSU rRNA regions confirmed the presence of the following fungal species in the samples analyzed: (1) Pestalotiopsis cocculi from the region I; (2) Epicoccum nigrum and Trichaptum biforme from the region I; (3) Nigrospora sphaerica and Fusarium sp. from the region II; and (4) Curvularia sp., Fusarium sp., Penicillium sp., and Preussia sp. from the region III. Conclusions: Our results suggest that the LSU and ITS regions are good genetic markers to perform fungal typing. Highlights: DNA sequencing technology can be used in the implementation of effective environmental monitoring programs of public health importance.

Genetic variability of wildlife-derived Sarcoptes scabiei determined by the ribosomal ITS-2 and mitochondrial 16S genes.

Infestation by the ectoparasitic mite Sarcoptes scabiei (Acari: Sarcoptidae) has important implications for global wildlife conservation and both animal and human health. Ribosomal and mitochondrial DNA sequences of parasites are useful to determine genetic diversity and to describe their likely dynamic evolution. In this study, we described the genetic diversity of S. scabiei individuals collected from wild animals in China by sequencing the ribosomal ITS-2 and mitochondrial 16S rRNA genes. A total of 13 Sarcoptes isolates of wildlife, coupled with one of rabbit origin, were subjected to genetic characteristics. After cloning and sequencing, 14 ITS-2 sequences and 12 16S rRNA sequences were obtained and analyzed. Further analysis of haplotype network and population genetic structure revealed that there were 79 haplotypes in ITS-2 (main haplotype H2) and 31 haplotypes in 16S rRNA (main haplotype C10). The phylogenetic trees showed some partial clustering by location and host, and the analysis of gene polymorphism may prompt that all isolates of S. scabiei have a similar origin. We speculate that the genetic evolution of S. scabiei may be related with that of the hosts, but more research is necessary to better understand the host-parasite co-evolutionary relationship in S. scabiei. These results provide new insights into understanding the population genetics and evolutionary biology of S. scabiei and therefore a better understanding of controlling its infestation pathways worldwide.

Bipolar dispersal of red-snow algae.

Red-snow algae are red-pigmented unicellular algae that appear seasonally on the surface of thawing snow worldwide. Here, we analyse the distribution patterns of snow algae sampled from glaciers and snow patches in the Arctic and Antarctica based on nuclear ITS2 sequences, which evolve rapidly. The number of phylotypes is limited in both polar regions, and most are specific to either the Arctic or Antarctica. However, the bipolar phylotypes account for the largest share (37.3%) of all sequences, suggesting that red-algal blooms in polar regions may comprise mainly cosmopolitan phylotypes but also include endemic organisms, which are distributed either in the Arctic or Antarctica.

Origin and diversification of winged bean (Psophocarpus tetragonolobus (L.) DC.), a multipurpose underutilized legume.

PREMISE OF THE STUDY: For many crops, research into the origin and partitioning of genetic variation is limited and this can slow or prevent crop improvement programs. Many of these underutilized crops have traits that could be of benefit in a changing climate due to stress tolerance or nutritional properties. Winged bean (Psophocarpus tetragonolobus (L.) DC.) is one such crop. All parts of the plant can be eaten, from the roots to the seeds, and is high in protein as well as other micronutrients. The goal of our study was to identify the wild progenitor and analyze the partitioning of genetic variation in the crop. METHODS: We used molecular phylogenetic analyses (cpDNA and nuclear ITS sequencing) to resolve relationships between all species in the genus, and population genetics (utilizing microsatellites) to identify genetic clusters of winged bean accessions and compare this to geography. KEY RESULTS: We find that winged bean is genetically distinct from all other members of the genus. We also provide support for four groups of species in the genus, largely, but not completely, corresponding to the results of previous morphological analyses. Within winged bean, population genetic analysis using 10 polymorphic microsatellite markers suggests four genetic groups; however, there is little correspondence between the genetic variation and the geography of the accessions. CONCLUSIONS: The true wild progenitor of winged bean remains unknown (or is extinct). There has likely been large-scale cross-breeding, trade, and transport of winged bean and/or multiple origins of the crop.

Examination of prezygotic and postzygotic isolating barriers in tropical Ulva (Ulvophyceae, Chlorophyta): evidence for ongoing speciation.

Phylogenetic clades based on DNA sequences such as the chloroplast rbcL gene and the nuclear ITS region are frequently used to delimit algal species. However, these molecular markers cannot accurately delimit boundaries among some Ulva species. Although Ulva reticulata and Ulva ohnoi occasionally bloom in tropical to warm-temperate regions and are clearly distinguishable by their reticulate or plain blade morphology, they have few or no sequence divergences in these molecular markers and form a monophyletic clade. In this study, to clarify the speciation and species delimitation in the U. reticulata-ohnoi complex clade, reproductive relationships among several sexual strains from the Philippines and Japan including offspring that originated from the type specimen of U. ohnoi were examined by culturing and hybridization in addition to the ITS-based analysis. As a result, both prezygotic and postzygotic reproductive isolation were revealed to occur between genetically perforated U. reticulata and imperforate U. ohnoi. They were also separated on the basis of sequence analysis of the ITS region. That strongly supports that the two taxa are independent biological species. Although no prezygotic barrier among the Philippine and Japanese strains of U. reticulata was observed, unexpectedly zoospores produced by hybrid sporophytes in some of their combinations mostly failed to develop, indicating partial formation of a postzygotic barrier despite a 0.2% divergence in the ITS sequence. These findings suggest speciation is still ongoing in U. reticulata.

Molecular Identification of Thrips Species Infesting Cotton in the Southeastern United States.

Traditional identification of thrips species based on morphology is difficult, laborious, and especially challenging for immature thrips. To support monitoring and management efforts of thrips as consistent and widespread pests of cotton (Gossypium hirsutum L.), a probe-based quantitative PCR (qPCR) assay with crude DNA extraction was developed to allow efficient and specific identification of the primary species of thrips infesting cotton. The assay was applied to identify over 5,000 specimens of thrips (including 3,366 immatures) collected on cotton seedlings from Alabama, Georgia, North Carolina, South Carolina, and Virginia in 2016. One half of all adult samples were examined by morphological identification, which provided a statistically equivalent species composition as the qPCR method. Frankliniella fusca (Hinds) (Thysanoptera: Thripidae) was the dominant species across all the locations (76.8-94.3% of adults and 81.6-98.0% of immatures), followed by Frankliniella occidentalis (Pergande) (Thysanoptera: Thripidae) in Georgia, North Carolina, and Virginia (4.6-19% of adults and 1.7-17.3% of immatures) or Frankliniella tritici (Fitch) (Thysanoptera: Thripidae) in South Carolina (10.8% of adults and 7.8% of immatures). Thrips tabaci (Lindeman) (Thysanoptera: Thripidae) and Neohydatothrips variabilis (Beach) (Thysanoptera: Thripidae) were occasionally found among adults but were rarely present among immature thrips. These five species of thrips represented 98.2-100% of samples collected across the Southeast. The qPCR assay was demonstrated to be a valuable tool for large-scale monitoring of species composition of thrips at different life stages in cotton. The tool will contribute to a better understanding of thrips population structure in cotton and could assist with development and application of improved management strategies.

Phylogenetic Evidence for the Existence of Multiple Strains of Rickettsia parkeri in the New World.

The bacterium Rickettsia parkeri has been reported to infect ticks of the "Amblyomma maculatum species complex" in the New World, where it causes spotted fever illness in humans. In South America, three additional rickettsial strains, namely, Atlantic rainforest, NOD, and Parvitarsum, have been isolated from the ticks Amblyomma ovale, Amblyomma nodosum, and Amblyomma parvitarsum, respectively. These three strains are phylogenetically closely related to R. parkeri, Rickettsia africae, and Rickettsia sibirica Herein, we performed a robust phylogenetic analysis encompassing 5 genes (gltA, ompA, virB4, dnaA, and dnaK) and 3 intergenic spacers (mppE-pur, rrl-rrf-ITS, and rpmE-tRNAfMet) from 41 rickettsial isolates, including different isolates of R. parkeri, R. africae, R. sibirica, Rickettsia conorii, and strains Atlantic rainforest, NOD, and Parvitarsum. In our phylogenetic analyses, all New World isolates grouped in a major clade distinct from the Old World Rickettsia species (R. conorii, R. sibirica, and R. africae). This New World clade was subdivided into the following 4 clades: the R. parkerisensu stricto clade, comprising the type strain Maculatum 20 and all other isolates of R. parkeri from North and South America, associated with ticks of the A. maculatum species complex; the strain NOD clade, comprising two South American isolates from A. nodosum ticks; the Parvitarsum clade, comprising two South American isolates from A. parvitarsum ticks; and the strain Atlantic rainforest clade, comprising six South American isolates from the A. ovale species complex (A. ovale or Amblyomma aureolatum). Under such evidences, we propose that strains Atlantic rainforest, NOD, and Parvitarsum are South American strains of R. parkeriIMPORTANCE Since the description of Rickettsia parkeri infecting ticks of the "Amblyomma maculatum species complex" and humans in the New World, three novel phylogenetic close-related rickettsial isolates were reported in South America. Herein, we provide genetic evidence that these novel isolates, namely, strains Atlantic rainforest, NOD, and Parvitarsum, are South American strains of R. parkeri. Interestingly, each of these R. parkeri strains seems to be primarily associated with a tick species group, namely, R. parkerisensu stricto with the "Amblyomma maculatum species group," R. parkeri strain NOD with Amblyomma nodosum, R. parkeri strain Parvitarsum with Amblyomma parvitarsum, and R. parkeri strain Atlantic rainforest with the "Amblyomma ovale species group." Such rickettsial strain-tick species specificity suggests a coevolution of each tick-strain association. Finally, because R. parkerisensu stricto and R. parkeri strain Atlantic rainforest are human pathogens, the potential of R. parkeri strains NOD and Parvitarsum to be human pathogens cannot be discarded.

Adding new pieces to the Azadinium (Dinophyceae) diversity and biogeography puzzle: Non-toxigenic Azadinium zhuanum sp. nov. from China, toxigenic A. poporum from the Mediterranean, and a non-toxigenic A. dalianense from the French Atlantic.

The marine planktonic dinophyceaen genus Azadinium is a primary source of azaspiracids, but due to their small size its diversity may be underestimated and information on its biogeography is still limited. A new Azadinium species, A. zhuanum was obtained from the East China Sea and Yellow Sea of China by incubating surface sediments. Five strains were established by isolating single germinated cells and their morphology was examined with light microscopy and scanning electron microscopy. Azadinium zhuanum was characterized by a plate pattern of Po, cp, X, 4', 2a, 6'', 6C, 5S, 6''', 2'''', by a distinct ventral pore at the junction of Po, the first and fourth apical plates, and a conspicuous antapical spine. Moreover, Azadinium poporum was obtained for the first time from the Mediterranean by incubating surface sediment collected from Diana Lagoon (Corsica) and a new strain of Azadinium dalianense was isolated from the French Atlantic. The morphology of both strains was examined. Small subunit ribosomal DNA (SSU rDNA), large subunit ribosomal DNA (LSU rDNA) and internal transcribed spacer (ITS) sequences were obtained from cultured strains. In addition, LSU sequences were obtained by single cell sequencing of two presumable A. poporum cells collected from the French Atlantic. Molecular phylogeny based on concatenated SSU, LSU and ITS sequences revealed that A. zhuanum was closest to A. polongum. French A. poporum from Corsica (Mediterranean) and from the Atlantic showed some genetic differences but were nested within one of the A. poporum ribotypes together with other European strains. Azadinium dalianense from France together with the type strain of the species from China comprised a well resolved clade now consisting of two ribotypes. Azaspiracid profiles were analyzed for the cultured Azadinium strains using LC-MS/MS and demonstrate that the Mediterranean A. poporum strain produced AZA-2 and AZA-2 phosphate with an amount of 0.44fgcell-1. Azadinium zhuanum and A. dalianense did not produce detectable AZA. Results of the present study support the view of a high diversity and wide distribution of species belonging to Azadinium. The first record of AZA-2 producing A. poporum from the Mediterranean suggests that this species may be responsible for azaspiracid contaminations in shellfish from the Mediterranean Sea.

Taxonomy and Phylogeny of Polyporus Group Melanopus (Polyporales, Basidiomycota) from China.

Melanopus is a morphological group of Polyporus which contains species with a black cuticle on the stipe. In this article, taxonomic and phylogenetic studies on Melanopus group were carried out on the basis of morphological characters and phylogenetic evidence of DNA sequences of multiple loci including the internal transcribed spacer (ITS) regions, the large subunit nuclear ribosomal RNA gene (nLSU), the small subunit nuclear ribosomal RNA gene (nSSU), the small subunit mitochondrial rRNA gene sequences (mtSSU), the translation elongation factor 1-α gene (EF1-α), the largest subunit of RNA polymerase II (RPB1), the second largest subunit of RNA polymerase II (RPB2), and ß-tubulin gene sequences (ß-tubulin). The phylogenetic result confirmed that the previously so-called Melanopus group is not a monophyletic assemblage, and species in this group distribute into two distinct clades: the Picipes clade and the Squamosus clade. Four new species of Picipes are described, and nine new combinations are proposed. A key to species of Picipes is provided.

Molecular diagnosis of Eimeria stiedae in hepatic tissue of experimentally infected rabbits.

The early detection of Eimeria stiedae in the hepatic tissue of experimentally infected rabbits was investigated using molecular assay. Forty 6-week-old male New Zealand rabbits were divided into two groups. Group A (30 animals) was infected with 2.5 × 10(4) sporulated oocysts of E. stiedae per animal on Day 0 and Group B (10 animals) was used as the uninfected controls. Three animals from Group A and one from Group B were sacrificed at 0, 3, 6, 9, 12, 15, 18, 21, 24 and 27 days post infection (PI). Gross and microscopic post-mortem findings were recorded. Polymerase chain reaction (PCR) of the E. stiedae internal transcribed spacer 1 genomic region was conducted on blood, liver tissue, and feces from the Group A experimentally infected animals. Macroscopically, the liver showed irregular yellowish white nodules pathognomonic to E. stiedae infection beginning on Day 15 PI. Hepatomegaly and ascites were obvious from Day 21-24 PI. The presence of different E. stiedae schizonts and gametocytes in the histopathological sections of the biliary epithelium were evident on Day 15 PI. The E. stiedae PCR was first positive in liver tissues on Day 12 and in fecal samples on Day 18 PI, but the blood samples were negative. In conclusion, the PCR can be used for early diagnosis and control of E. stiedae schizonts before shedding of the oocysts in feces.

Isolation and identification of Malassezia species from Chinese and Korean patients with seborrheic dermatitis and in vitro studies on their bioactivity on sebaceous lipids and IL-8 production.

We investigated the distribution of Malassezia yeast in 120 Chinese (20 patients from each of six cities) and 20 Korean patients with scalp seborrheic dermatitis (SD) and dandruff (SD/D) using ITS1 and ITS2 polymerase chain reaction-restriction fragment length polymorphism. Bioactivity was studied by quantifying sebum lipid production by human primary sebocytes and inflammatory cytokine, interleukin-8 (IL-8) production was studied by exposing HaCaT keratinocytes with extracts of five standard Malassezia strains; M. globosa, M. restricta, M. sympodialis, M. dermatis and M. slooffiae. M. restricta and M. globosa were the most frequently encountered species from both Chinese and Korean patients. These two Malassezia species also promoted neutral lipid synthesis although the result was not statistically significant and induced significant increase in IL-8 production among the five Malassezia species studied. The study suggests a possible role of these organisms in the pathogenesis of SD/D.

Identification of maca (Lepidium meyenii Walp.) and its adulterants by a DNA-barcoding approach based on the ITS sequence.

Maca (Lepidium meyenii) is an herbaceous plant that grows in high plateaus and has been used as both food and folk medicine for centuries because of its benefits to human health. In the present study, ITS (internal transcribed spacer) sequences of forty-three maca samples, collected from different regions or vendors, were amplified and analyzed. The ITS sequences of nineteen potential adulterants of maca were also collected and analyzed. The results indicated that the ITS sequence of maca was consistent in all samples and unique when compared with its adulterants. Therefore, this DNA-barcoding approach based on the ITS sequence can be used for the molecular identification of maca and its adulterants.