Molecular characterization of 39 de novo sSMC: contribution to prognosis and genetic counselling, a prospective study.

Small supernumerary marker chromosomes (sSMCs) are structurally abnormal chromosomes that cannot be characterized by karyotype. In many prenatal cases of de novo sSMC, the outcome of pregnancy is difficult to predict because the euchromatin content is unclear. This study aimed to determine the presence or absence of euchromatin material of 39 de novo prenatally ascertained sSMC by array-comparative genomic hybridization (array-CGH) or single nucleotide polymorphism (SNP) array. Cases were prospectively ascertained from the study of 65,000 prenatal samples [0.060%; 95% confidence interval (CI), 0.042-0.082]. Array-CGH showed that 22 markers were derived from non-acrocentric markers (56.4%) and 7 from acrocentic markers (18%). The 10 additional cases remained unidentified (25.6%), but 7 of 10 could be further identified using fluorescence in situ hybridization; 69% of de novo sSMC contained euchromatin material, 95.4% of which for non-acrocentric markers. Some sSMC containing euchromatin had a normal phenotype (31% for non-acrocentric and 75% for acrocentric markers). Statistical differences between normal and abnormal phenotypes were shown for the size of the euchromatin material (more or less than 1 Mb, p = 0.0006) and number of genes (more or less than 10, p = 0.0009). This study is the largest to date and shows the utility of array-CGH or SNP array in the detection and characterization of de novo sSMC in a prenatal context.

Wyeomyia exallos, a new species of sylvatic mosquito (Diptera: Culicidae) from Brazil

Wyeomyia exallos, a new mosquito species from Atlantic Forest of southeastern Brazil, is described based on morphological characters of the adult female, male, male genitalia, pupa and fourth-instar larva. The morphological characters of Wy. exallos sp. nov. are compared with those of different subgenera of Wyeomyia as well as of species without subgeneric position. It is proposed that the new species should be placed in genus Wyeomyia Theobald without subgeneric assignment.

Clinic-epidemiologic study of human infection by Granada virus, a new phlebovirus within the sandfly fever Naples serocomplex.

Granada virus (GRV), a new phlebovirus within the Naples serocomplex, has been recently described in phlebotomine sandflies from Spain. The presence of anti-GRV immunoglobulin G (IgG) antibodies was investigated by indirect fluorescence assay (IFA) and neutralization test (NT) in 920 serum samples from the Granada population. By IFA, an overall GRV seroprevalence of 15.8% (N = 145) was observed, significantly increasing up to 65 years. NT was positive in 18% of anti-GRV IFA-positive samples. IgG antibodies against Toscana virus (TOSV), a hyperendemic phlebovirus within Granada province, were detected in 40% of anti-GRV-positive cases. Anti-GRV IgM antibodies were detected in 36 (6.6%) of 547 acute-phase serum samples from individuals with febrile illness, exanthema, and/or acute respiratory infection. All positives were anti-TOSV IgM-negative. GRV may infect humans, with most cases being asymptomatic. The codetection of anti-GRV and anti-TOSV IgG antibodies could be attributable to cross-reactivity or exposure to the same transmission vector.

A male with unilateral microphthalmia reveals a role for TMX3 in eye development.

Anophthalmia and microphthalmia are important birth defects, but their pathogenesis remains incompletely understood. We studied a patient with severe unilateral microphthalmia who had a 2.7 Mb deletion at chromosome 18q22.1 that was inherited from his mother. In-situ hybridization showed that one of the deleted genes, TMX3, was expressed in the retinal neuroepithelium and lens epithelium in the developing murine eye. We re-sequenced TMX3 in 162 patients with anophthalmia or microphthalmia, and found two missense substitutions in unrelated patients: c.116G>A, predicting p.Arg39Gln, in a male with unilateral microphthalmia and retinal coloboma, and c.322G>A, predicting p.Asp108Asn, in a female with unilateral microphthalmia and severe micrognathia. We used two antisense morpholinos targeted against the zebrafish TMX3 orthologue, zgc:110025, to examine the effects of reduced gene expression in eye development. We noted that the morphant larvae resulting from both morpholinos had significantly smaller eye sizes and reduced labeling with islet-1 antibody directed against retinal ganglion cells at 2 days post fertilization. Co-injection of human wild type TMX3 mRNA rescued the small eye phenotype obtained with both morpholinos, whereas co-injection of human TMX3(p.Arg39Gln) mutant mRNA, analogous to the mutation in the patient with microphthalmia and coloboma, did not rescue the small eye phenotype. Our results show that haploinsufficiency for TMX3 results in a small eye phenotype and represents a novel genetic cause of microphthalmia and coloboma. Future experiments to determine if other thioredoxins are important in eye morphogenesis and to clarify the mechanism of function of TMX3 in eye development are warranted.

Ecologia del tram final del riu Ebre: la comunitat de macròfits i macroinvertebrats associats

Durant els últims anys al tram final del riu Ebre s’han produit canvis molt importants a l’ecosistema fluvial: l’augment de la transparència de l’aigua ha comportat una proliferació massiva de macròfits que ha provocat canvis en l’estructura tròfica i en la composició de les comunitats biològiques, representant un greu perill per espècies amenaçades com Margaritifera auricularia. A més del problema ecològic, els macròfits estan provocant molts problemes socio-econòmics perjudicant les captacions d’aigua (centrals nuclears, hidroelèctriques i regadius), creant problemes per a la navegació fluvial, i afavorint la proliferació d’espècies molestes com la mosca negra (Simulium erythrocephalum). Entre les diferents causes que podrien explicar aquests canvis en l’ecosistema hi ha: la disminució del fòsfor dissolt, la regularització i la disminució de cabals, i l’aparició d’espècies introduides com el musclo zebra. Segurament es tractarà d’un efecte combinat de les diferents causes però és necessari analitzar-les per tal de conèixer quines tenen més incidència i així, poder proposar mesures de gestió per als problemes ecològics que pateix el tram final de l’Ebre. Al present projecte de tesi (inclós en el projecte d’I+D: efectes de la millora de la qualitat de l'aigua i de l'alteració del règim de cabals sobre les comunitats biològiques del tram final del riu Ebre) s’estudiarà la comunitat de macròfits i macroinvertebrats associats per tal de determinar el paper que tenen en el canvis que s’han produit al riu durant els últims anys.

Kazal-type serine proteinase inhibitors in the midgut of Phlebotomus papatasi

Sandflies (Diptera: Psychodidae) are important disease vectors of parasites of the genus Leishmania, as well as bacteria and viruses. Following studies of the midgut transcriptome of Phlebotomus papatasi, the principal vector of Leishmania major, two non-classical Kazal-type serine proteinase inhibitors were identified (PpKzl1 and PpKzl2). Analyses of expression profiles indicated that PpKzl1 and PpKzl2 transcripts are both regulated by blood-feeding in the midgut of P. papatasi and are also expressed in males, larva and pupa. We expressed a recombinant PpKzl2 in a mammalian expression system (CHO-S free style cells) that was applied to in vitro studies to assess serine proteinase inhibition. Recombinant PpKzl2 inhibited α-chymotrypsin to 9.4% residual activity and also inhibited α-thrombin and trypsin to 33.5% and 63.9% residual activity, suggesting that native PpKzl2 is an active serine proteinase inhibitor and likely involved in regulating digestive enzymes in the midgut. Early stages of Leishmania are susceptible to killing by digestive proteinases in the sandfly midgut. Thus, characterising serine proteinase inhibitors may provide new targets and strategies to prevent transmission of Leishmania.

Effect of Temperature on the Development and Survival of the Argentine Ant, Linepithema humile

The influence of temperature on the developmental times and survival of insects can largely determine their distribution. For invasive species, like the Argentine ant, Linepithema humile Mayr (Hymenoptera: Formicidae), these data are essential for predicting their potential range based on mechanistic models. In the case of this species, such data are too scarce and incomplete to make accurate predictions based on its physiological needs. This research provides comprehensive new data about brood survival and developmental times at a wide range of temperatures under laboratory conditions. Temperature affected both the complete brood development from egg to adult worker and each of the immature stages separately. The higher the temperature, the shorter the development times. Brood survival from egg to adult was low, with the maximum survival rate being only 16% at 26° C. Temperature also affected survival of each of the immature stages differently: eggs were negatively affected by high temperatures, while larvae were negatively affected by low temperatures, and the survival of pupae was apparently independent of environmental temperature. At 32° C no eggs survived, while at 18° C less than 2% of the eggs hatched into larva. The data from the present study are essential for developing prediction models about the distribution range of this tramp species based on its physiological needs in relation to temperature

Coupled reproductive ecologies in a plant-pollinator-seed predator system

Résumé de la thèseBien que le mutualisme puisse être considéré comme une relation harmonieuse entre différentes espèces, son étude révèle plutôt une exploitation réciproque où chaque partenaire tente de maximiser ses bénéfices tout en réduisant ses coûts. Dans ce contexte, l'identification des facteurs qui favorisent ou contrarient, au cours de l'évolution, une issue mutualiste est une étape majeure pour pouvoir reconstruire les étapes clés menant à l'apparition et au maintien des interactions mutualistes. Le but de ce doctorat était l'identification des traits phénotypiques qui permettent à la plante Silene latofolia (Caryophyllacée)et à son pollinisateur - prédateur de graines, la phalène Hadena bicruris (Noctuidé), d'augmenter les bénéfices nets que chacun retire de l'interaction. Ce système d'étude est particulièrement bien approprié à l'étude de ces traits, car on peut assez facilement estimer la qualité et la quantité des descendants (fitness) des deux partenaires. En effet, la femelle papillon pond un oeuf dans la fleur qu'elle pollinise et sa larve se développe dans le fruit, consommant les graines de la plante. Ainsi, sur une même plante, il est possible d'estimer les succès respectifs de la plante et du papillon à obtenir une descendance. De plus, le conflit d'intérêt autour des graines qui sont indispensables, à la fois à la plante et au papillon, peut stimuler l'évolution de traits qui limitent la surexploitation réciproque des partenaires. Dans une première étude, j'ai montré que le papillon mâle était un pollinisateur efficace de S. latifolia et qu'ainsi, il permettait à la plante d'augmenter le nombre de graines produites (i.e.bénéfice) sans pour autant augmenter la quantité de larves sur la plante. Dans ce système, les papillons pondent un seul oeuf par fleur, déposé soit à l'intérieur de la fleur, dans le tube de corolle, soit sur le pétale. Ma seconde étude montre que les plantes répondent différemment à la présence des oeufs suivant leur position. Aussi, quand l'oeuf est placé dans la fleur, la plante a davantage tendance à ne pas développer le fruit de la fleur infesté ou bien à produire des fruits plus petits que lorsque l'oeuf est placé sur le pétale. Enfin, j'ai montré que la femelle du papillon pond plus souvent sur le pétale lorsque elle visite des fleurs dotées d'un long tube de corolle, et que les larves issues de ces oeufs ont moins de chances de réussir à pénétrer dans le fruit que les larves issues des oeufs placés à l'intérieur de la fleur. Aussi, la variation observée du site de ponte pourrait être causé par la morphologie de la fleur qui contraint le papillon à pondre sur le pétale. Vu dans leur ensemble, les résultats obtenus pendant ce doctorat suggèrent que la participation des mâles à la pollination, l'absence de développement des fruits et la profondeur du tube de corolle pourraient réduire les coûts que S. latifolia subit dans son interaction avec H. bicruris. Par ailleurs, je n'ai pas détecté de mécanismes qui permettraient au papillon de réduire les coûts que la plante pourrait lui imposer. La prochaine étape serait de déterminer l'effet des traits identifiés dans ce doctorat sur la fitness globale de la plante et du papillon pour estimer pleinement leur efficacité à réduire les coûts et à favoriser une issue mutualiste. De même, il faudrait évaluer l'effet de ces traits en populations naturelles pour identifier le rôle des facteurs environnementaux sur leur efficacité.AbstractAlthough mutualisms can be regarded as harmonious relationships between the interacting partners, they are best conceptualized as reciprocal exploitations in which each partner attempts to increase its own benefits and decrease its costs. To date, identifying the factors which promote or discourage mutualistic outcomes remains a major goal to reconstruct the ecological conditions leading to mutualisms. The aim of this PhD thesis was to identify phenotypic traits that may increase the net benefits of each partner in the interaction between the plant Silene latifolia (Caryophyllaceae) and its pollinator / seed predator, the moth Hadena bicruris (Noctuidae). This study system is particularly well suited because the fitness of both interacting species can be assessed. The female moth lays its egg in the flower it pollinated, and its offspring grows in the fruit, feeding on the seeds of the plant, which allows for the follow-up of both larva and fruit fates. Furthermore, the inherent conflict of interest over the seeds as plant progeny vs. larval resource may stimulate the evolution of traits that reduce overexploitation in both the moth and plant. In a first study, I show that male moths are efficient pollinators, hence increasing seed production without increasing oviposition. The contribution of male moths to pollination might thus improve the net benefits of the interaction for the host plant. Females of the H. bicruris moth lay a single egg per flower, and place it either inside the corolla tube or on the petal. My second study shows that plants are more likely to abort the infested flower or to produce a smaller fruit when the egg was experimentally placed inside the flower compared to plants that received an egg on the petal. Finally, female moths were found to lay their eggs more frequently on the petal when visiting a flower with a deep corolla tube, and larvae hatching from these eggs less likely to successfully attack the fruit. Variation in egg position on the flower may thus be the result of a constraint imposed by floral morphology. Overall, this PhD work suggests that the pollination by male moths, flower abortion, and deep corolla tube may efficiently reduce the costs experienced by S. latifolia in its interaction with H. bicruris. Interestingly, no apparent mechanism of costs reduction was detected for the moth. Further studies should focus on the effects of these traits (i) in the long term fitness of both the plant and the insect and (ii) their interactions with environmental factors (biotic and abiotic) that may affect their efficiency in natural populations.

Control of brood male production in the Argentine ant Iridomyrmex humilis (Mayr)

The influence of various social factors on the production of males was investigated in the Argentine ant, Iridomyrmex humilis. In this polygynous species, the workers which are monomorphic are unable to lay reproductive eggs, so all the males are the progeny of the queens. Although male eggs appear to be laid by mated queens throughout the year, in large stock colonies males are reared periodically (every 3 or 4 months); males develop from brood taken from these colonies at any point in the cycle and given queenless or queenright (1 to 5 queens) units. This is in striking contrast to many other species of ants where it is generally assumed that male eggs are laid seasonnally. Comparative experiments suggest that several related factors influence the rearing of males as far as the pupal stage. Worker/larva ratio: The proportion of male larvae developing in standardized units in which the worker/larva ratio was varied from 0.25 to 25 demonstrated that low ratios inhibit male production. Queen influence: In standardized units where the worker/larva ratio was high the presence of queens did not inhibit the rearing of males suggesting that there is no queen inhibitory pheromone controlling male experimental production. Data suggest evidence that queens prevent male production by means of appropriation of food. Diet: Male larvae failed to pupate in experimental societies deprived of protein. Thus, the production of males appears to be controlled by the amount of food available to larvae. This depends on foraging activity, the quantity of brood in relation to the number of workers and the number of queens in the society.

Systematic molecular and cytogenetic screening of 100 patients with marfanoid syndromes and intellectual disability.

The association of marfanoid habitus (MH) and intellectual disability (ID) has been reported in the literature, with overlapping presentations and genetic heterogeneity. A hundred patients (71 males and 29 females) with a MH and ID were recruited. Custom-designed 244K array-CGH (Agilent®; Agilent Technologies Inc., Santa Clara, CA) and MED12, ZDHHC9, UPF3B, FBN1, TGFBR1 and TGFBR2 sequencing analyses were performed. Eighty patients could be classified as isolated MH and ID: 12 chromosomal imbalances, 1 FBN1 mutation and 1 possibly pathogenic MED12 mutation were found (17%). Twenty patients could be classified as ID with other extra-skeletal features of the Marfan syndrome (MFS) spectrum: 4 pathogenic FBN1 mutations and 4 chromosomal imbalances were found (2 patients with both FBN1 mutation and chromosomal rearrangement) (29%). These results suggest either that there are more loci with genes yet to be discovered or that MH can also be a relatively non-specific feature of patients with ID. The search for aortic complications is mandatory even if MH is associated with ID since FBN1 mutations or rearrangements were found in some patients. The excess of males is in favour of the involvement of other X-linked genes. Although it was impossible to make a diagnosis in 80% of patients, these results will improve genetic counselling in families.

Programa de investigação sobre o ictioplancton em Cabo Verde

O conhecimento da história natural e das fases do ciclo de vida dos peixes ajuda-nos a conhecer e entender melhor a dinâmica ecológica da comunidade íctica. Estudos de distribuição e abundância do ictioplancton são importantes na determinação dos períodos e locais de desova. A identificação precisa desses locais tem importância fundamental na implementação de medidas de gestão, visando a gestão sustentável das espécies enquanto recurso marinho explorável. O crescimento e a mortalidade de peixes marinhos nos estágios iniciais dos ciclos de vida são muito variáveis. Pequenas variações em taxas de crescimento provocam grandes variações na sobrevivência e consequentemente, na disponibilidade do recurso para a pesca. Isto deve-se às diferentes mecanismos e interacções entre os factores ecológicos que influenciam a dinâmica dos gâmetas, ovos, larvas e juvenis de peixes. Acredita-se pois que tal seja uma das causas fundamentais que explica a variabilidade encontrada no recrutamento de recursos à pesca. A maior mortalidade dos peixes ocorre nos primeiros 100 dias de vida, durante o estado de ovo, larva e póslarva. Em 2002/2003, foi realizada, no canal entre São Vicente e Santo Antão, a primeira campanha especificamente dirigida ao ictioplancton na região. Os resultados obtidos e sua importância no desenvolvimento da pesca, motivaram a elaboração de um programa de estudo em Cabo Verde, que se prevê com uma duração inicial de 5 anos.

Contribuição para o estudo de bactrocera invadens em cabo verde

Bactrocera invadens (Drew, Tsuruta & White) (Diptera:Tephritidae) é uma das espécies de moscas-das-frutas de maior importância económica para a fruticultura mundial. De origem asiática, surgiu em África em 2003, tendo sido detectada em Cabo Verde, Ilha de Santiago, em 2007. Desde então, espalhou-se para Santo Antão, Fogo e Brava. Tendo em vista a avaliação da eficiência de armadilhas para captura em massa da mosca, procedeu-se, durante curtos períodos de tempo em 2009, 2010 e 2011, à recolha de adultos com recurso a dois tipos de atractivos, ‘Creolax’® e proteína hidrolisada, e a três tipos de armadilhas, ABT®, armadilha do tipo funil e garrafa mosqueira da CeraTrap®. As armadilhas foram instaladas em quatro localidades de Santiago, na cultura de mangueira ou de bananeira. O ‘Creolax’ foi significativamente superior à proteína hidrolisada e a armadilha ABT apresentou elevada eficiência na captura de adultos da mosca. Num ensaio em 2009, na Cidade Velha, foram capturados 195, 59 e 30 adultos/armadilha para as combinações, respectivamente, de ABT x ‘Creolax’, armadilha do tipo funil x ‘Creolax’ e garrafa mosqueira x proteína hidrolisada. Em 2011, na bananeira, em Monte Negro, foram capturados 8500 adultos/armadilha ABT x ‘Creolax’ em cinco meses, tendo o máximo da população sido encontrado em Junho.

Maternal effect on female caste determination in a social insect.

Caste differentiation and division of labor are the hallmarks of social insect colonies [1, 2]. The current dogma for female caste differentiation is that female eggs are totipotent, with morphological and physiological differences between queens and workers stemming from a developmental switch during the larval stage controlled by nutritional and other environmental factors (e.g., [3-8]). In this study, we tested whether maternal effects influence caste differentiation in Pogonomyrmex harvester ants. By conducting crossfostering experiments we identified two key factors in the process of caste determination. New queens were produced only from eggs laid by queens exposed to cold. Moreover, there was a strong age effect, with development into queens occurring only in eggs laid by queens that were at least two years old. Biochemical analyses further revealed that the level of ecdysteroids was significantly lower in eggs developing into queens than workers. By contrast, we found no significant effect of colony size or worker exposure to cold, suggesting that the trigger for caste differentiation may be independent of the quantity and quality of resources provided to larvae. Altogether these data demonstrate that the developmental fate of female brood is strongly influenced by maternal effects in ants of the genus Pogonomyrmex.

Leaf consumption and duration of instars of the cassava defoliator Erinnyis ello (L., 1758) (Lepidoptera, Sphingidae)

The objectives of this research were to evaluate leaf consumption and the developmental time of the larvae of Erynnyis ello (L., 1758) (Lepidoptera, Sphingidae) reared on cassava, in order to obtain information for the integrated management of this pest. The larvae were reared on excised cassava leaves in Petri dishes and later in gerbox, and kept in chambers at 24 ± 2 ºC and 75 ± 10% RH. The total leaf area consumed by the larva to complete its development was 589.67 cm²; each of the five instars consumed, respectively: 1.89 cm²; 5.74 cm²; 17.48 cm²; 76.66 cm²; and 487.90 cm². The consumption by the first three instars was insignificant, and did not reach 5% altogether; the 4th represented 13%; the 5th presented a consumption significantly higher, about 82.7%. The total time for the larval development was 22.61 days, and the duration for each of the five larval instar was, respectively: 4.35; 3.19; 3.32; 4.52; and 4.94 days. The pre-pupal period lasted 2.29 days. Since the highest consumption is by the 5th instar larva, the control should be applied before this age to avoid heavier damages to the cassava crop.

Morfologia externa dos estágios imaturos de heliconíneos neotropicais: I. Eueides isabella dianasa (Hübner, 1806)

External morphology of the immature stages of neotropical heliconians: I. Eueides isabella dianasa (Hübner, 1806). The external features of egg, larva and pupa of Eueides isabella dianasa (Hübner, 1806) (Lepidoptera, Nymphalidae, Heliconiinae) are described and illustrated, based upon light and scanning electron microscopy.