Genus-specific kinetoplast-DNA PCR and parasite culture for the diagnosis of localised cutaneous leishmaniasis: applications for clinical trials under field conditions in Brazil

The positivities of two methods for the diagnosis of localised cutaneous leishmaniasis (CL) were estimated in 280 patients enrolled in a clinical trial. The trial was conducted in an endemic area of Leishmania (Viannia) braziliensis and trial participants were patients with skin ulcers and positive leishmanin skin tests. Patients underwent aspirative skin punctures of the ulcerated lesions and lymph nodes for in vitro cultures, which were processed under field conditions at the local health centre. Skin lesion biopsies were tested at a reference laboratory using kinetoplastid DNA (kDNA)-PCR to detect DNA. The median time required to obtain a positive culture from the skin samples was seven days and the contamination rate of the samples was 1.8%. The positivities of the cultures from skin lesions, kDNA-PCR and the combination of the two methods were 78.2% (95% CI: 73-82.6%), 89.3% (95% CI: 85.1-92.4%) and 97.1% (95% CI: 94.5-98.5%). We conclude that parasite culture is a feasible method for the detection of Leishmania in field conditions and that the combination of culture and PCR has a potential role for the diagnosis of CL in candidates for clinical trials.

Distribution and habitat preferences of the genus Biomphalaria (Gastropoda: Planorbidae) in Cuba

A study was carried out to determine the distribution and habitat preferences of several species of the genus Biomphalaria. Samples were taken at 350 freshwater locations in Cuba. Three species of Biomphalaria (Biomphalaria havanensis, Biomphalaria helophila and Biomphalaria pallida) were recorded based on their distribution. Of the three species, B. havanensis has the weakest distribution because it was identified in only one locality. The other species, B. helophila and B. pallida, are abundant in rivers and dams and have large populations in Cuba. However, the only species that appears to occur in ecosystems shared with thiarids is B. pallida, possibly due to recent introduction of thiarids, but always in fewer numbers. Here we discuss the possibility of these species to act as intermediary hosts of Schistosoma mansoni in Cuba over the basis of occurrence in natural and anthropic habitats.

Biological and genetic aspects of crosses between species of the genus Meccus (Hemiptera: Reduviidae Triatominae)

The degree of reproductive isolation between Meccus phyllosomus and the remaining five species of the genus Meccus, as well as between Meccus bassolsae and Meccus pallidipennis, Meccus longipennis and Meccus picturatus, was examined. Fertility and the segregation of morphological characteristics were examined in two generations of hybrids from crosses between these species. The percentage of couples with offspring (fertile) was high in the vast majority of sets of crosses, with the exception of that between ♀M. phyllosomus and ♂Meccus mazzottii. In sets of crosses involving M. bassolsae specimens, no first-generation (F1) individuals were morphologically similar to M. bassolsae, but instead shared the morphology of the other parental species. A similar phenomenon was observed in most sets of crosses involving M. phyllosomus. These results indicated that different degrees of reproductive isolation exist among the species of Meccus involved in this study. The biological evidence obtained in this study does not support the proposal that M. bassolsae is a full species. It could indicate that, on the contrary, it should be considered a subspecies of a single polytypic species. The biological evidence does support the proposal that M. phyllosomus is a full species.

Description of the cave organ in three species of the genus Belminus (Hemiptera: Reduviidae: Triatominae) by optical and scanning electron microscopy

The cave organ is a sensory receptor in the antenna pedicel of some Reduviidae. This paper describes this organ for the first time in three species of the genus Belminus, Belminus corredori, Belminus ferroae and Belminus herreri, by optical and scanning electron microscopy. The structures presented a general pattern similar to one reported for other species of Triatominae.

DNA multigene sequencing of topotypic specimens of the fascioliasis vector Lymnaea diaphana and phylogenetic analysis of the genus Pectinidens (Gastropoda)

Freshwater lymnaeid snails are crucial in defining transmission and epidemiology of fascioliasis. In South America, human endemic areas are related to high altitudes in Andean regions. The species Lymnaea diaphana has, however, been involved in low altitude areas of Chile, Argentina and Peru where human infection also occurs. Complete nuclear ribosomal DNA 18S, internal transcribed spacer (ITS)-2 and ITS-1 and fragments of mitochondrial DNA 16S and cytochrome c oxidase (cox)1 genes of L. diaphana specimens from its type locality offered 1,848, 495, 520, 424 and 672 bp long sequences. Comparisons with New and Old World Galba/Fossaria, Palaearctic stagnicolines, Nearctic stagnicolines, Old World Radix and Pseudosuccinea allowed to conclude that (i) L. diaphana shows sequences very different from all other lymnaeids, (ii) each marker allows its differentiation, except cox1 amino acid sequence, and (iii) L. diaphana is not a fossarine lymnaeid, but rather an archaic relict form derived from the oldest North American stagnicoline ancestors. Phylogeny and large genetic distances support the genus Pectinidens as the first stagnicoline representative in the southern hemisphere, including colonization of extreme world regions, as most southern Patagonia, long time ago. The phylogenetic link of L. diaphana with the stagnicoline group may give light to the aforementioned peculiar low altitude epidemiological scenario of fascioliasis.

The phylogeny of Simulium (Chirostilbia) (Diptera: Simuliidae) and perspectives on the systematics of the genus in the Neotropical Region

Recently, knowledge of Neotropical Simuliidae has been accumulating quickly. However, information about supra-specific relationships is scarce and diagnoses of Simulium subgenera are unsatisfactory. To investigate the relationships among Simulium (Chirostilbia) species and test the subgenus monophyly, we performed a cladistic analysis. The ingroup included all species of this subgenus and the outgroup included representatives of the 17 species groups of Neotropical Simulium and three Holarctic species. The study was based on a data matrix with 31 terminal taxa and 45 morphological characteristics of adult, pupa and larva. The phylogenetic analysis under equal weights resulted in eight most-parsimonious trees (length = 178, consistency index = 34, retention index = 67). The monophyly of the S. (Chirostilbia) was not supported in our analysis. The Simulium subpallidum species group was closer to Simulium (Psilopelmia) and Simulium (Ectemnaspis) than to the Simulium pertinax species group. Additionally, we describe the three-dimensional shape of the terminalia of male and female of Simulium (Chirostilbia) for the first time and provide comments about the taxonomic problems involving some species of the subgenus: Simulium acarayense, Simulium papaveroi, S. pertinax, Simulium serranum, Simulium striginotum and S. subpallidum.

Genetic and morphological characterisation of a new species of the genus Hysterothylacium (Nematoda) from Paralichthys isosceles Jordan, 1890 (Pisces: Teleostei) of the Neotropical Region, state of Rio de Janeiro, Brazil

Taking into account the difficulties of taxonomic identification of larval anisakid nematodes based on morphological characters, genetic analyses were performed, together with those usually applied, in order to identify anisakid larvae found in the flounder Paralichthys isosceles from the littoral of the state of Rio de Janeiro, Brazil. The analysis of 1,820 larvae revealed a new species, similar to Hysterothylacium MD, Hysterothylacium 2, Hysterothylacium KB and Hysterothylacium sp regarding the absence of the larval tooth, an excretory pore situated below the nerve ring level, and slender lateral alae. Moreover, the new species differs from Hysterothylacium fortalezae and Hysterothylacium reliquens with regard to the number and size of spines present on the tail end and from Hysterothylacium patagonicus by the absence of interlabia. The maximum parsimony and neighbour joining tree topologies based on the 18S ribosomal DNA gene, complete internal transcribed spacer region and cytochrome oxidase 2 (COII) gene demonstrated that the Brazilian larvae belong to Raphidascarididae and represent a unique genetic entity, confirmed as a new Hysterothylacium species. Furthermore, the new species presents COII genetic signatures and shares polymorphisms with Raphidascarididae members. This is the first description of a new anisakid species from Brazil through the integration of morphological and molecular taxonomy data.

1.° Guillelmi Phicheti tractatus de punctis, virgulis, aliisque id genus. — 2.° Tricenarium pro adolescentibus, sive triginta regulae grammaticae, quae videntur esse ejusdem Phicheti.

Baluzianus

Two new species and new records of biting midges of the genus Culicoides from northwestern Argentina (Diptera: Ceratopogonidae)

The following two new species of Culicoides from the Argentinean Yungas are described, illustrated and placed to subgenus or species group and compared with related congeners: Culicoides calchaqui Spinelli & Veggiani Aybar and Culicoides willinki Spinelli & Veggiani Aybar. Culicoides daedaloides Wirth & Blanton is recorded for the first time for Argentina and Culicoides pseudoheliconiae Felippe-Bauer is firstly mentioned from the northwestern region of the country.

Revision of the genus acrochordiceras hyatt, 1877 (Ammonoidea, Middle Triassic): Morphology, biometry, biostratigraphy and intra-specific variability

The family Acrochordiceratidae Arthaber, 1911 ranges in age from latest Spathian to the middle/late Anisian boundary, and it represents a major component of ammonoid faunas during that time. The middle Anisian genus Acrochordiceras Hyatt, 1877 is the most widespread taxon of the family and occurs abundantly worldwide within the low paleolatitude belt. However, there is a profusion of species names available for Acrochordiceras. This excessive diversity at the species level essentially results from the fact that sufficiently large samples were not available, thus leading to a typological approach to its taxonomy. Based on new extensive collections obtained from the Anisian (Middle Triassic) Fossil Hill Member (Star Peak Group, north-west Nevada) for which a high resolution biostratigraphic frame is available, the taxonomy and biostratigraphy of the genus Acrochordiceras Hyatt, 1877 is herein revised with respect to its intra-specific variation. Morphological and biometric studies (c. 550 bedrock-controlled specimens were measured) show that only one species occurs in each stratigraphic level. Continuous ranges of intra-specific variation of studied specimens enable us to synonymize Haydenites Diener, 1907, Silesiacrochordiceras Diener, 1916 and Epacrochordiceras Spath, 1934 with Acrochordiceras Hyatt, 1877. Three stratigraphically successive species are herein recognized in the low paleolatitude middle Anisian faunas from Nevada: A. hatschekii (Diener, 1907), A. hyatti Meek, 1877 and A. carolinae Mojsisovics, 1882. Moreover, an assessment of intra-specific variation of the adult size range does not support recognition of a dimorphic pair (Acrochordiceras and Epacrochordiceras) as previously suggested by other workers (Epacrochordiceras is the compressed and weakly ornamented end-member variant of Acrochordiceras). The successive middle Anisian species of Acrochordiceras form an anagenetic lineage characterized by increasing involution, adult size and intra-specific variation. This taxonomic revision based on new bedrock-controlled collections is thus an important prerequisite before studying the evolution of the group.

A Comparative study of germination strategies of two species of genus Allium sect. Allium

Germination experiments were performed with seeds of two species of genus Allium section Allium, a rare and endangered species A. pyrenaicum and a common A. sphaerocephalon. Different pre-treatments and a photoperiod of 24 h darkness were applied in order to simulate different germination conditions. Both species showed a high percentage of viable seeds a part of which were dormant. An elevate percentage of dormant seeds could be caused by a later collection time. Low altitude populations had more mortality than the others, possibly caused by the hard summer conditions during flowering and fruiting time. Comparisons between dates of species coexistence localities only show inter-population variability and it could be caused by the detected dormancy. Darkness accelerates germination, possibly for elongation radicle stimulation. Heat-shock pre-treatments decreased germination time in seeds from localities where fire is a probable event. The rarity of A. Pyrenaicum not seems to be caused by restricted germination requirements but is attributable to distinct habitat preferences, related to his altitudinal range of distribution

Genetic structure of the genus Leptospira by multilocus enzyme electrophoresis

Thirty strains from the 11 species of the genus Leptospira were studied by multilocus enzyme electrophoresis at 12 enzyme loci, all of which were polymorphic. The mean number of alleles per locus was 6.5. Twenty-five electrophoretic types were distinguished. Grouping of the strains by cluster analysis was in general agreement with species delineation as determined by DNA-DNA hybridization, except for the strains of Leptospira meyeri and Leptospira inadai, which were scattered throughout the genus, reflecting previously recognized taxonomic uncertainties. Analysis of the clonality within Leptospira interrogans sensu stricto indicated that this population was relatively heterogeneous and a lack of gene linkage disequilibrium could not be excluded. There was a genetic discrimination between the pathogenic species and the saprophytic ones. The phenotypically intermediate species (L. inadai and Leptospira fainei) were also genetically separated and were probably closer to the saprophytes than to the pathogens.

Low dispersing species : population genetic processes in space and time : the genus "Trochulus" (Gastropoda: Hygromiidae) as a case study

RÉSUMÉ Une espèce est rarement composée d'une population unique. Parce que les individus ont des capacités de dispersion limitées et que les paysages sont des mosaïques d'habitats, la plupart des espèces sont plutôt composées de sous-populations connectées par la migration. Cette variation spatiale influence directement la distribution de la variabilité génétique dans et entre les populations. Durant ce travail, nous avons abordé certains des processus populationnels qui ont joué un rôle supposé dans l'apparition de nouvelles espèces au sein du genre Trochulus. Plus précisément, nous avons tenté d'évaluer les impacts respectifs de l'isolement passé (facteurs historiques) et présent (facteurs locaux). Nous avons d'abord pu montrer que les faibles capacités de dispersion des escargots terrestres ont directement influencé leur histoire évolutive à toutes les échelles spatiales et temporelles. En réduisant l'effet homogénéisant de la migration, une faible dispersion maintient dans les populations les traces génétiques d'évènements passés. A l'échelle de la distribution globale de Trochulus villosus, ces traces ont permis de reconstruire une histoire faite d'isolements et d'expansions de populations. En combinant des données génétiques avec une modélisation de la niche climatique passée, il a été possible de proposer un scénario significativement meilleur que toutes les hypothèses alternatives que nous avons testées. A l'échelle locale par contre, l'héritage historique est difficile à distinguer de la dynamique actuelle. Ce fut le cas des lignées mitochondriales du complexe sericeus-hispidus : les deux principales lignées étaient phylogénétiquement éloignées, avaient eu des démographies passées différentes et corrélaient avec des différences morphologiques. D'un autre côté, le flux de gène nucléaire était fort, contredisant l'idée de deux espèces cryptiques isolées reproductivement. Pour pouvoir conclure à la présence ou non de deux espèces, il nous a manqué des informations locales sur la dynamique des populations et les conditions écologiques que l'on trouve dans la région d'étude. Enfin, nous avons pu souligner que la connectivité entre populations d'escargots est soumise à la qualité des habitats et à leur organisation spatiale. Les escargots sont dépendants d'un habitat et s'y adaptent, comme l'indiquent la présence de «poils » uniquement sur la coquille d'espèces vivant dans des habitats humides ou la corrélation entre morphologie et habitat au sein du complexe sericeus-hispidus. Logiquement donc, les escargots migrent préférentiellement au travers d'habitats favorables comme l'a montré la réduction de flux de gènes au travers des prairies chez T. villosus (une espèce forestière). De ces données, nous pouvons supposer que les populations d'escargots en particulier, et des espèces à faible dispersion en général, ont de fortes chances d'être affectées par les changements climatiques, avec de probables implications pour leurs histoires évolutives. SUMMARY : Species rarely consists in a single population. Because individuals have limited dispersal abilities, because landscapes are habitat patchworks, most species are made of several subpopulations connected by migration. This spatial variation has consequences on the distribution of genetic diversity within and between populations, creating a structure among the populations. During the present work, we investigated some of the population processes assumed to have played an important role on the speciation within the genus Trochulus. More specifically, we questioned the respective impacts of past (historical factors) or present (local factors) population isolations. We first could show that the poor dispersal abilities of land snails have had profound impacts on their evolutionary histories at all spatial and temporal scales. Low dispersal maintains a strong signature of past events in the populations by minimising the homogenising effects of geneflow. At the scale of Trochulus villosus global distribution, they allowed to retrieve the detailed history of this species population isolations and expansions. Combining a large genetic dataset with paleo-climatic niche modelling ended up with a historical scenario significantly better than all traditional alternatives we tested. At local scale on the contrary, past events become difficult to tease apart from ongoing processes. This was the case for the divergent mitochondria) lineages within the sericeus-hispidus complex: the two principal lineages appeared to be phylogenetically distant, to have experienced different demographic histories and to correlate with morphological differences. On the other hand, nuclear (present day) geneflow was high, contradicting the idea of two reproductively isolated cryptic species. Information on the local population dynamics and environmental conditions are lacking to be able to decide whether past isolation has indeed resulted here in new species. Finally, we emphasised the importance of the habitat types present in a landscape as well as their spatial organisation for the population connectivity of land snails. These species are tightly dependent on a habitat and adapt to it as shown by thé occurrence of hair-like structures only in species living in humid environments or by the correlation between shell morphology and habitat in the sericeus-hispidus complex. As a result, land snails preferentially migrate through favourable habitats: Trochulus villosus, a forest species, had its geneflow significantly reduced across meadows. From these data, we can hypothesise that the populations of land snails in particular and of low dispersing species in general are likely to be strongly affected by the ongoing climate changes, with potential major consequences on their evolutionary histories.

Graeciae Antiquae Tabula Nova in qua locorum situs tum ad distantias itinerarias tum ad Observationes Astronomicas exactus litorum flexurae et alia id genus ad accuratas recentiorum rationes accomodatae su[n]t. Graeciae Pars Meridionalis / Autore Guillelmo Delisle, ...

Échelle(s) : [1:1 275 000 environ], Scala, Stadia Graeca quorum 580. gradui respondent [= 8,7 cm]

Graeciae Antiquae Tabula Nova in qua locorum situs tum ad distantias itinerarias tum ad Observationes Astronomicas exactus litorum flexurae et alia id genus ad accuratas recentiorum rationes accomodatae su[n]t. Graeciae Pars Septentrionalis / Autore Guillelmo Delisle, ...

Échelle(s) : [1:1 275 000 environ], Scala, Stadia Graeca quorum 580. gradui respondent [= 8,7 cm]