Molecular phylogeny of black flies (Diptera: Simuliidae) from Thailand, using ITS2 rDNA

The sequences of the second internal transcribed spacer (ITS2) of ribosomal DNA (rDNA) were determined for 40 black fly species from Thailand, belonging to 4 subgenera of the genus Simulium, namely Gomphostilbia (12 species), Nevermannia (5 species), Montisimulium (1 species), Simulium sensu stricto (21 species), and an unknown subgenus with one species (Simulium baimaii). The length of the ITS2 ranged from 247 to 308 bp. All black fly species had high AT content, ranging from 71 to 83.8%. Intraindividual variation (clonal variation) occurred in 13 species, ranging from 0.3 to 1.1%. Large intrapopulation and interpopulation heterogeneities exist in S. feuerboni from the same and different locations in Doi Inthanon National Park, northern Thailand. Phylogenetic relationships among 40 black fly species were examined using PAUP (version 4.0b10) and MrBAYS (version 3.0B4). The topology of the trees revealed two major monophyletic clades. The subgenus Simulium and Simulium baimaii were placed in the first monophyletic clade, whereas the subgenera Nevermannia + Montisimulium were placed as the sister group to the subgenus Gomphostilbia in the second monophyletic clade. Our results suggest that S. baimaii belongs to the malyschevi-group or variegatum-group in the subgenus Simulium. The molecular phylogeny generally agrees with existing morphology-based phylogenies.     

西藏蚋属一新种（双翅目：蚋科）

记述西藏自治区蚋属Simulium蚋亚属Simulium 1新种,以陈汉彬教授的名命名为汉彬蚋Simulium(Simulium) hanbini sp. nov.。该新种与Simulium (Simulium) jingfui Cai & An, 2008及Simulium (Simulium)liubaense Liu & An, 2009相似,但该蚋爪无基齿,拉氏器较长,食窦的形状也不同。模式标本保存在北京军事医学科学院医学昆虫标本馆。     

Polytene chromosome maps in four species of tsetse flies Glossina austeni, G. pallidipes, G. morsitans morsitans and G. m. submorsitans (Diptera: Glossinidae): a comparative analysis

Photographic polytene chromosome maps from pupal trichogen cells of four tsetse species, Glossina austeni, G. pallidipes, G. morsitans morsitans and G. m. submorsitans were constructed and compared. The homology of chromosomal elements between the species was achieved by comparing banding patterns. The telomeric and subtelomeric chromosome regions were found to be identical in all species. The pericentromeric regions were found to be similar in the X chromosome and the left arm of L1 chromosome (L1L) but different in L2 chromosome and the right arm of L1 chromosome (L1R). The L2 chromosome differs by a pericentric inversion that is fixed in the three species, G. pallidipes, G. morsitans morsitans and G. m. submorsitans. Moreover, the two morsitans subspecies appeared to be homosequential and differ only by two paracentric inversions on XL and L2L arm. Although a degree of similarity was observed across the homologous chromosomes in the four species, the relative position of specific chromosome regions was different due to chromosome inversions established during their phylogeny. However, there are regions that show no apparent homology between the species, an observation that may be attributed to the considerable intra—chromosomal rearrangements that have occurred following the species divergence. The results of this comparative analysis support the current phylogenetic relationships of the genus Glossina.     

Phylogenetic relationships among the black fly species (Diptera: Simuliidae) of Thailand based on multiple gene sequences

Simulium is a very speciose genus of the black fly family Simuliidae that includes many important pests of humans and animals. Cytotaxonomic and morphological studies have made substantial progress in Simulium systematics. 16S rRNA and ITS-1 DNA sequence studies have assisted this progress. Intensive multi-gene molecular systematic investigations will, however, be required for a comprehensive understanding of the genus’ taxonomy and evolution. Our research was conducted to investigate the relationships of Thai Simulium at the subgeneric, species group and species levels. We also examined the possibility of using mitochondrial DNA sequences to facilitate Simulium species identification. Data were collected from three mitochondrial genes (COI, ND4 and 16S rRNA) and two segments of the nuclear 28S ribosomal RNA (the D1 to D2 and the D4 expansion regions). The subgenera Simulium and Gomphostilbia were monophyletic in most analyses. Nevermannia included Montisimulium but was otherwise monophyletic in multigene analyses. In most analyses, Simulium and Nevermannia were more closely related to each other than to Gomphostilbia which was usually basal. Species groups were generally monophyletic. Within Gomphostilbia, however, the batoense species group was always paraphyletic to the other two species groups found in Thailand. Three species groups in Simulium were not monophyletic. The tendency to gill filament number reduction for some species groups in the subgenus Simulium was associated with a derived position in multigene analyses. Most species were monophyletic with two exceptions that probably represent species complexes and will present difficulties for rapid mitochondrial DNA identification.     

A multi‐locus approach resolves the phylogenetic relationships of the Simulium asakoae and Simulium ceylonicum species groups in Malaysia: evidence for distinct evolutionary lineages

A multi‐locus approach was used to examine the DNA sequences of 10 nominal species of blackfly in the Simulium subgenus Gomphostilbia (Diptera: Simuliidae) in Malaysia. Molecular data were acquired from partial DNA sequences of the mitochondria‐encoded cytochrome c oxidase subunit I (COI), 12S rRNA and 16S rRNA genes, and the nuclear‐encoded 18S rRNA and 28S rRNA genes. No single gene, nor the concatenated gene set, resolved all species or all relationships. However, all morphologically established species were supported by at least one gene. The multi‐locus sequence analysis revealed two distinct evolutionary lineages, conforming to the morphotaxonomically recognized Simulium asakoae and Simulium ceylonicum species groups.     

Two new species and one new variety of Oudemansiella (Agaricales) from Japan

Two new species and one new variety of Oudemansiella (Agaricales) from Japan are described and illustrated: (1) Oudemansiella latilamellata sp. nov. (subgenus Xerula, section Radicatae) has relatively broad lamellae and broadly ellipsoid spores with a subacute apex; (2) Oudemansiella rhodophylla sp. nov. (subgenus Xerula, section Radicatae), growing in deciduous forests of Fagus crenata, Quercus crispula, etc., is characteristic in having lamellae tinted reddish; and (3) Oudemansiella amygdaliformis var. bispora var. nov. (subgenus Xerula, section Albotomentosae) is distinguished from the type variety by having two-spored basidia and grows in evergreen oak forests (Castanopsis sieboldii, Quercus glauca, etc.) or bamboo groves.     

A new black fly species of the subgenus Gomphostilbia (Simulium: Simuliidae: Diptera) from India

Simulium (Gomphostilbia) agasthyamalaiense sp. nov. is described based on adults, pupae and mature larvae from a medium-flowing stream of Southern Western Ghats, India. This new species is placed in the Simulium batoense species-group of the subgenus Gomphostilbia Enderlein. This new species is characterized in the female by a scutum with three brownish-black longitudinal vittae and the hind basitarsus 5.7 times as long as wide; in the female by the large facets of upper eye with 20 vertical columns and 19 horizontal rows; in the pupa the respiratory gill with medium-long common basal stalk; and in the larva arrowhead-shaped postgenal cleft. Taxonomic notes are provided for this new species and it can be distinguished from closely related species of S. (G.) peteri. Keys are constructed to distinguish this species from ten species of the batoense species-group recorded in India.www.zoobank.org/urn:lsid:zoobank.org:pub:1D1CFEE5-A762-4F68-9EAE-9E31133146C0     

The genomic relationship between Glycine max (L.) Merr. and G. soja Sieb. and Zucc. as revealed by pachytene chromosome analysis

Summary This study was conducted with the objective of determining the genomic relationship between cultivated soybean (Glycine max) and wild soybean (G. soja) of the subgenus Soja, genus Glycine. Observations on cross-ability rate, hybrid viability, meiotic chromosome pairing, and pollen fertility in F ₁ hybrids of G. max × G. soja and reciprocals elucidated that both species hybridized readily and set mature putative hybrid pods, generated vigorous F₁ plants, had a majority of sporocytes that showed 18II + 1IV chromosome association at diakinesis and metaphase I, and had a pollen fertility that ranged from 49.2% to 53.3%. A quadrivalent was often associated with the nucleolus, suggesting that one of the chromosomes involved in the interchange is a satellited chromosome. Thus, G. max and G. soja genetic stocks used in this study have been differentiated by a reciprocal translocation. Pachytene analysis of F₁ hybrids helped construct chromosome maps based on chromosome length and euchromatin and heterochromatin distribution. Chromosomes were numbered in descending order of 1–20. Pachytene chromosomes in soybean showed heterochromatin distribution on either side of the centromeres. Pachytene analysis revealed small structural differences for chromosomes 6 and 11 which were not detected at diakinesis and metaphase I. This study suggests that G. max and G. soja carry similar genomes and validates the previously assigned genome symbol GG.Research supported in part by the Illinois Agricultural Experiment Station and U.S. Department of Agriculture Competitive Research Grant (85-CRCR-1-1616)     

四照花亚属的nrDNA ITS致同进化不完全

四照花亚属(Cornus subg.Syncarpea)隶属于山茱萸科山茱萸属(Cornus),我国该亚属共有5种8亚种。为探讨四照花亚属nrDNA ITS序列的致同进化不完全现象及假基因产生的可能原因,分析了该亚属4种(每种1~2个居群)共21个个体的nrDNA ITS序列。结果表明,这些类群的nrDNA ITS存在多态性,通过分析这些nrDNA ITS克隆序列的G+C含量、5.8S保守基序和二级结构最小自由能,推测其可能存在假基因。系统发育研究结果显示所有nrDNA ITS序列分成5个分支,同一个体的不同拷贝被分别置于两个甚至多个分支中,且不同分支显示了不同种间关系。四照花亚属物种个体内部存在nrDNA ITS不完全致同进化,可能归咎于不完全的世系分选(incomplete lineage sorting)、种间杂交或多倍化等进化事件,从而导致基因组内nrITS区序列出现多态性,同时也导致难以通过外部形态来划分亚属内种间界限。     

Comparative karyotype analysis in Haplopappus Cass. and Grindelia Willd. (Asteraceae) by double FISH with rRNA specific genes

The genera Grindelia Willd. and Haplopappus Cass. belong to the family Asteraceae - Astereae and are distributed in America and South America, respectively. Previous cytotaxonomic studies showed for South American species of Grindelia 2n=12 and for Haplopappus 2n=10 and 2n=12. Both Grindelia species (G. anethifolia, G. prunelloides), newly analyzed with molecular-cytological methods, exhibited symmetric karyotypes (AsI %=55.46 and 55.95) with metacentric chromosome sets (5m + 1m-sat) and 2n=12 chromosomes. The NOR was detected after fluorescence in situ hybridization (FISH) with 18/25S rDNA in the satellite chromosome 2. In contrast H. Happlopappus glutinosus, H. grindeloides and H. stolpii showed exclusively a higher asymmetric index (66.83%, 67.01% and 68.87%, respectively) with submetacentric chromosome sets (4sm + 1sm–sat). The sat-chromosomes 3 of H. glutinosus and H. grindelioides were both significantly different in their length from chromosomes 2 and 4. Furthermore in Grindelia the FISH with 5S rDNA could estimate signals in the short arms of chromosomes 3 or 4, that were not significantly differentiated in their length. Contrary to these findings in Grindelia, the position of 5S rDNA in Haplopappus was detected in the long arms of chromosome 1 (H. grindelioides and H. stolpii) and chromosome 2 (with two different loci) and chromosome 4 of H. glutinosus. The lengths of all measured chromosome arms with 5S rDNA were significantly different to those of the neighbours in the karyotypes. The two-color FISH of 5S and 18/25S rDNA had provided clear karyotypic markers for three (Haplopappus glutinosus) and two (H. grindelioides and H. stolpii) chromosomes. The number and position of rDNA signals were relatively highly conserved in the investigated five species without the double marked chromosome 2 of H. glutinosus, which shows an evolutionary dynamic of this 5S rRNA specific gene cluster. This investigation supports the assumption that the evolution of New World members of Grindelia and Haplopappus has not been accompanied by large karyotypic changes, but small chromosomal rearrangements have undoubtedly occurred (e.g. 5S rDNA localizations).     

Characterization of Charr Chromosomes Using Fluorescence in Situ Hybridization

The chromosomal locations of several families of tandem repetitive DNA sequences and the 5S rDNA were determined using fluorescence in situ hybridization (FISH) in the five North American charr species: Salvelinus namaycush, S. fontinalis, S. alpinus, S. malma, and S. confluentus. The pattern of hybridization of three centromeric repetitive sequences previously isolated from S. namaycush and S. alpinus was unique in each species. Dual-color FISH experiments showed that in several species many of the centromeres had the EcoRI-DraI family in addition to either the AluI-RsaI type A or type B families. The EcoRI-DraI family which was found only at the centromeres of acrocentric chromosomes in S. namaycush, S. fontinalis and S. malma was also found at centromeres of selected metacentrics in S. alpinus (one pair) and S. confluentus (four pairs) whose chromosomes have undergone additional centric fusions compared to the other species. The locations of 5S rDNA sequences were different in each species except for the two most closely related (S. alpinus and S. malma). Two whole-arm chromosome paint probes, one specific for the short and the other for the long arm of the lake charr sex chromosomes, hybridize to the same chromosome pair in all species. Results with other paint probes suggest that independent centric fusions have occurred in S. alpinus and S. confluentus which is consistent with the phylogenetic tree obtained previously for Salvelinus with cytogenetic and DNA data.     

Marker chromosomes commonly observed in the genus Glycine

Summary Soybean [Glycine max (L.) Merr.] chromosomes were analyzed using the chromosome image analyzing system, CHIAS, and seven groups, including subgroups, were identified based on morphological characteristics. Two pairs of chromosomes were conspicuous in their morphological traits. One pair of chromosomes, which had the largest arm ratio among all the chromosomes, was commonly observed in the species in all three subgenera of the genus Glycine. These chromosomes also displayed a unique pattern after N-banding and were detected as marker chromosomes. G. soja, which is considered to be the ancestor of G. max, has two types of marker chromosomes. The lines that carry the same type as G. max may be the ancestors of G. max among the lines of G. soja. The morphological differences of the marker chromosomes within the species in the subgenus Soja are discussed in relation to the domestication process of soybean.     

A simple chromosomal marker can reliably distinguishes <Emphasis Type="Italic">Poncirus</Emphasis> from <Emphasis Type="Italic">Citrus</Emphasis> species

Several chromosome types have been recognized in Citrus and related genera by chromomycin A₃ (CMA) banding patterns and fluorescent in situ hybridization (FISH). They can be used to characterize cultivars and species or as markers in hybridization and backcrossing experiments. In the present work, characterization of six cultivars of P. trifoliata (“Barnes”, “Fawcett”, “Flying Dragon”, “Pomeroy”, “Rubidoux”, “USDA”) and one P. trifoliata × C. limonia hybrid was performed by sequential analyses of CMA banding and FISH using 5S and 45S rDNA as probes. All six cultivars showed a similar CMA⁺ banding pattern with the karyotype formula 4B + 8D + 6F. The capital letters indicate chromosomal types: B, a chromosome with one telomeric and one proximal band; D, with only one telomeric band; F, without bands. In situ hybridization labeling was also similar among cultivars. Three chromosome pairs displayed a closely linked set of 5S and 45S rDNA sites, two of them co-located with the proximal band of the B type chromosomes (B/5S-45S) and the third one co-located with the terminal band of a D pair (D/5S-45S). The B/5S-45S chromosome has never been found in any citrus accessions investigated so far. Therefore, this B chromosome can be used as a marker to recognize the intergeneric Poncirus × Citrus hybrids. The intergeneric hybrid analyzed here displayed the karyotype formula 4B + 8D + 6F, with two chromosome types B/5S-45S and two D/5S-45S. The karyotype formula and the presence of two B/5S-45S chromosomes clearly indicate that the plant investigated is a symmetric hybrid. It also demonstrates the suitability of karyotype analyses to differentiate zygotic embryos or somatic cell fusions involving trifoliate orange germplasm. During the submission of this paper, we analyzed 25 other citrus cultivars with the same methodology and we found that the chromosome marker reported here can indeed distinguish Poncirus trifoliata from grapefruits, pummelos, and one variegated access of Citrus, besides the previously reported access of limes, limons, citrons, and sweet-oranges. However, among 14 mandarin cultivars, two of them displayed a single B/5S-45S chromosome, whereas in Citrus hystrix D.C., a far related species belonging to the Papeda subgenus, this chromosome type was found in homozygosis. Since these two mandarin cultivars are probably of hybrid origin, we assume that for almost all commercial cultivars and species of the subgenus Citrus this B type chromosome is a useful genetic marker.     

Cytosystematics of the Simulium tuberosum group (Diptera: Simuliidae) in Thailand

The polytene chromosomes of 3347 larvae of the Simulium tuberosum group in Asia were analysed, representing the largest ever cytogenetic study of black flies in the Oriental Region. Band‐by‐band comparisons, relative to the established standard chromosome map for the subgenus Simulium, revealed 17 cytogenetically distinct taxa in Thailand, plus an 18th in China. Six of these taxa correspond to morphologically described species (S. doipuiense, S. rufibasis, S. setsukoae, S. tani, S. yuphae and S. weji). Recognition of the 18 taxa is based largely on unique inversions, either fixed or sex linked, primarily in the long arm of chromosome III. The greatest cytological diversity was discovered in the S. tani lineage, with ten cytoforms. This marked chromosomal diversification within S. tani is based largely on two inversions that have assumed different roles over evolutionary time, variously functioning in different combinations as fixed inversions, sex‐linked inversions and autosomal polymorphisms. Shared unique chromosomal features, relative to the subgeneric standard chromosome map, allowed evolutionary relationships among the cytotaxa to be inferred. Fluctuations in climate during the Pleistocene might have promoted differentiation of the Southeast Asian S. tuberosum group in isolated refugia such as mountains. © 2009 The Linnean Society of London, Zoological Journal of the Linnean Society, 2009, 155 , 289–315.     

Characterization of geographically isolated accessions in five Alstroemeria L. species (Chile) using FISH of tandemly repeated DNA sequences and RAPD analysis

In fifteen geographically isolated populations of five species of Alstroemeria L. (A. aurea, A. hookeri, A. ligtu, A. pelegrina and A. presliana) collected in Chile, karyotypes and variation of RAPD markers were investigated. Tandemly repeated DNA sequences - 5S and 18/25S rDNA genes and the sequence A001-1 (De Jeu et al. 1997) were used to characterize karyotypes by fluorescence in situ hybridization (FISH). Ten somatic metaphases per population were used for measurement of chromosome length. Differences in RAPD marker bands were used for characterization of populations, creating a similarity index. FISH with all three DNA probes shows a high degree of polymorphism between and sometimes also within accessions of A. aurea, A. hookeri and A. ligtu. The number of chromosome pairs showing 5S rDNA signals is more different for the investigated species A. aurea, A. hookeri, A. ligtu, A. pelegrina and A. presliana with 5, 7, 5, 3 and 7, respectively, than the number of 18/25S rDNA signals in this succession with 7, 7, 6, 5 and 7 chromosome pairs, showing a high evolutionary dynamics within the genus. Furthermore, among the four populations of A. hookeri, accession 4181 was different in arm length of chromosome 3. RAPD markers (index of similarity) also showed a greater genetic distance of accession 4181 from the other three accessions of A. hookeri. The possible evolutionary mechanisms providing these polymorphisms were discussed.     

Three new West Palaearctic species of Rhamphomyia subgenus Lundstroemiella (Diptera, Empididae)

Rhamphomyia (Lundstroemiella) cervi sp. n. (France), R. (L.) cimrmani sp. n. (Turkey) and R. (L.) speighti sp. n. (France) are described and illustrated. A key to the Palaearctic species of the subgenus Lundstroemiella Frey is given.     

Cytogenetics of Simulium siamense Takaoka and Suzuki, 1984 (Diptera: Simuliidae) in northeastern Thailand

We analysed salivary gland polytene chromosomes of 796 larvae from 17 populations of Simulium siamense in northeastern Thailand. Seventeen floating and two fixed chromosome inversions were recorded. Three cytoforms (A, F and G) were recognised and two of them are new (F and G). Cytoform F is distinguished by a fixed inversion on the long arm of chromosome II (IIL-8) and cytoform G by fixed inversions on the long arm of chromosome II (IIL-8) and short arm of chromosome III (IIIS-2). Significant departures from Hardy–Weinberg equilibrium due to heterozygote deficiency in geographically intermediate populations and absence of shared polymorphic inversions of the cytoforms indicate separation of the gene pool. Morphometric analysis of the larvae revealed significant differences in body length (F = 5.00, p =0.007) and head capsule width (F = 4.68, p = 0.010) among cytoforms.     

New species and records of saprophytic ascomycetes (Myxotrichaceae) from decaying logs in the boreal forest

Decayed wood from fallen white spruce (Picea glauca) and trembling aspen (Populus tremuloides) collected in northeastern Alberta, Canada, was the source of new isolates of species in the ascomycete generaGymnostellatospora andPseudogymnoascus. In addition to new reports ofG. japonica, G. frigida andP. roseus, two new species are described.Gymnostellatospora canadensis sp. nov. resemblesG. japonica but differs in producing brown ascomata and in the formation of an arthroconidial anamorph.Gymnostellatospora subnuda sp. nov. is distinct in lacking differentiated peridial hyphae.Gymnostellatospora alpina was not found in decayed wood but is reviewed based on extralimital material. A dichtomous key to the five species ofGymnostellatospora is provided.     

Molecular phylogeny and systematics of Genista (Leguminosae) and related genera based on nucleotide sequences of nrDNA (ITS region) and cpDNA (trnL-trnF intergenic spacer)

Phylogenetic relationships of Genista and related genera (Teline, Chamaespartium, Pterospartum, Echinospartum, Ulex, Stauracanthus and Retama) were assessed by the analysis of sequences of the nrDNA internal transcribed spacer (ITS region), and the cpDNA trnL-trnF intergenic spacer. The tree obtained by combining both sets of data indicates the existence of three lines of diversification within Genista, that correspond to three subgenera: Genista, Phyllobotrys and Spartocarpus, however, each of these lineages encompass also species of the related genera Echinospartum, Teline, Retama, Chamaespartium, Pterospartum, Ulex, Stauracanthus. The molecular data do not support division of these subgenera into taxonomical units at the sectional level; only sections Genista and Spartocarpus are monophyletic groups. The sequences of both regions are also informative at the specific level, grouping morphologically related species (e.g. the G. cinerea aggregate). The molecular data have also helped to clarify the position of taxa whose relationships were not well established (e.g. G. valdes-bermejoi). The relationships of related genera that belong to the Genista lines of diversification have also been investigated. Echinospartum splits into two separate clades matching the separation of two ecological and caryological differentiated groups. Teline also forms two groups, both placed near to Genista subgenus Genista, but that separated from the main core of the group. Retama, morphologically well differentiated from Genista, is close to Genista subgenus Spartocarpus. Chamaespartium and Pterospartum do not form a monophyletic group. Chamaespartium is closer to Genista subgenus Genista, whereas Pterospartum stands close to: 1) Genista subgenus Spartocarpus (particularly, sect. Cephalospartum); and 2) the Ulex-Stauracanthus clade (a terminal derivative of Genista subgenus Spartocarpus). Cases of incongruence (e.g. Echinospartum, Chamaespartium, Teline) between the trees obtained from the two molecular markers, may be indicating hybridisation and/or introgression between different lines of Genisteae.