Molecular and cellular analysis of genodermatoses

Summary Skin is the essential interface between our body and its environment; not only does it prevent water loss and protect us from external insults it also plays an essential role in the central nervous system acting as a major sense organ primarily for touch and pain. The main cell type present in skin, keratinocyte, undergoes a differentiation process leading to the formation of this protecting barrier. This work is intended to contribute to the understanding of how keratinocyte differentiates and skin functions. To do this, we studied two genetic skin diseases: Erythrokeratodermia variabilis and Mal de Meleda. Our approach was to examine the expression and localization of proteins implicated in these two pathologies in normal and diseased tissues and to determine the influence of mutant proteins at the molecular and cellular levels. Connexins are major components of gap junctions, channels allowing direct communication between cells. Our laboratory has identified mutations in both connexin 30.3 (Cx30.3) and 31 (Cx31) to be causally involved in erythrokeratodermia variabilis (EKV), an autosomal dominant disorder of keratinization. In the first chapter, we show a new mutation of Cx31, L209P-Cx31, in 3 EKV patients, extending the field of EKV-causing mutations although the mechanism by which connexin mutations lead to the disease is unclear. In the second chapter, we studied the effect of F137L-Cx30.3 on expression, trafficking and localization of cotransfected Cx31 and Cx30.3 in connexin-deficient HeLa cells. The F137 amino acid, highly conserved in connexin family, is oriented towards the channel pore and F137L mutation in either Cx30.3 or Cx31 lead to EKV. As two genes can lead to EKV when mutated, our hypothesis was that Cx31 and Cx30.3 might cooperate at a molecular level. We were able to demonstrate a physical interaction between Cx31 and Cx30.3. The presence of F137L-Cx30.3 disturbed the trafficking of both connexins, less connexins were integrated into gap junctions and thus, the coupling between cell was diminished. Connexins formed in the presence of F137L-Cx30.3 are degraded at their exit from the endoplasmic reticulum. In conclusion, our results indicate that the genetic heterogeneity of EKV is due to mutations in two interacting proteins. F137L-Cx30.3 has a dominant negative effect and affects Cx31, disturbing cellular communication in epidermal cells. Mal de Meleda is an autosomal recessive inflammatory and a keratotic palmoplantar skin disorder due to mutations in SLURP1 (secreted LY6/PLAUR-related protein 1). SLURP1 belongs to the LY6/PLAUR family of proteins and has the particularity of being secreted instead of being GPI-anchored. The high degree of structural similarity between SLURP1 and the three fingers motif of snake neurotoxins and LYNX 1-C suggests that this protein could interact with the neuronal acetylcholine receptors. In the third chapter, we show that SLURP1 potentiates responses of the a7 nicotinic acetylcholine receptor (nAchR) to acetylcholine. These results identify SLURP1 as a secreted epidermal neuromodulator that is likely to be essential for palmoplantar skin. In the fourth chapter, we show that SLURP1 is expressed in the granular layer of the epidermis but is absent from skin biopsies of Mal de Meleda patients. SLURP1 is also present in secretions such as sweat, tears or saliva. An in vitro analysis on two mutant of SLURP-I demonstrates that W15R-SLURP1 is absent in cells while G86R-SLURP1 is expressed and secreted, suggesting that SLURP1 can lead to the disease by either an absent or an abnormal protein. Finally, in the fifth chapter, we analyse the expression and biological properties of other LY6/PLAUR members, clustered around SLURP] on chromosome 8. Their GPI-anchored or secreted status were analysed in vitro. SLURP1, LYNX1-A and -B are secreted while LYPDC2 and LYNX 1-C are GPI anchored. Three of these proteins are expressed in the epidermis and in cultured keratinocytes. These results suggest that these LY6/PLAUR members may have an important role in skin homeostasis. Résumé Résumé La peau est la barrière essentielle entre notre corps et l'environnement, nous protégeant des agressions extérieures, de la déshydratation et assurant aussi un rôle dans le système nerveux central en tant qu'organe du toucher et de la douleur. Le principal type de cellules présent dans la peau est le kératinocyte qui suit un processus de différenciation aboutissant à la formation de cette barrière protectrice. Ce travail est destiné à comprendre la différenciation des kératinocytes et le fonctionnement de la peau. Pour cela, nous avons étudié deux maladies génodermatoses : l'Erthrokeratodermia Variabilis (EKV) et le Mal de Meleda. Nous avons examiné l'expression et la localisation des protéines impliquées dans ces deux pathologies dans des tissus normaux et malades puis déterminé l'influence des protéines mutantes aux niveaux moléculaires et cellulaires. Les connexines (Cx) sont les composants majeurs des jonctions communicantes, canaux permettant la communication directe entre les cellules. Notre laboratoire a identifié des mutations dans les Cx30.3 et Cx31 comme responsables de l'EKV, génodermatose de transmission autosomique dominante. Dans le ler chapitre, nous décrivons une nouvelle mutation de Cx31, L209-Cx31, et contribuons à l'établissement du catalogue des mutations de Cx31 entraînant cette maladie. Cependant, le mécanisme par lequel les mutations de Cx31 et C3x0.3 provoquent l'EKV est inconnu. Dans le 2ème chapitre, nous étudions les effets de la mutation F137L-Cx30.3 sur l'expression, le trafic et la localisation des Cx31 et Cx30.3 transfectées dans des cellules HeLa, déficientes en connexines. Comme deux gènes peuvent causer une EKV quand ils sont mutés, notre hypothèse était que Cx31 et Cx30.3 pourraient coopérer au niveau moléculaire. Nous avons montré l'existence d'une interaction physique entre ces deux connexines. La présence de la mutation F137L-Cx30.3 perturbe le trafic des deux connexines, moins de connexines sont intégrées dans les jonctions communicantes et donc le couplage entre les cellules est diminué. Les connexons formés en présence de cette mutation sont dégradés à leur sortie du réticulum endoplasmique. En conclusion, nos résultats indiquent que l'hétérogénéité génétique de EKV est due à des mutations dans deux protéines qui interagissent. F137L-Cx30.3 a un effet dominant négatif et affecte Cx31, perturbant la communication entre les cellules épidermiques. Le Mal de Meleda est une maladie récessive de la peau palmoplantaire due à des mutations dans SLURP1. SLURP1 appartient à la famille des protéines contenant un domaine LY6/PLAUR et a la particularité d'être sécrétée. La grande homologie de structure existant entre SLURP1, les neurotoxines de serpent et LYNX1-C suggère que la protéine pourrait interagir avec des récepteurs à acétylcholine (Ach). Dans le 3ème chapitre, nous montrons que SLURP1 module la réponse à l'Ach du récepteur nicotinique α7. Ces résultats identifient SLURP1 comme un neuromodulateur épidermique sécrété, probablement essentiel pour la peau palmoplantaire. Dans le 4ème chapitre, nous montrons que SLURP1 est exprimé dans la couche granuleuse de l'épiderme et qu'il est absent des biopsies des patients. SLURP1 a aussi été détecté dans des sécrétions telles que la sueur, les lamies et la salive. Une analyse in vitro de deux mutants de SLURP1 a montré que W15R-SLURP1 est absent des cellules tandis que G86R-SLURP1 est exprimé et sécrété, suggérant qu'une absence ou une anomalie de SLURP1 peuvent causer la maladie. Finalement, dans le 5ème chapitre, nous analysons l'expression et les propriétés biologiques d'autres membres de la famille LY6/PLAUR localisés autour de SLURP1 sur le chromosome 8. Leur statut de protéines sécrétées ou liées à la membrane par une ancre GPI est analysé in vitro. SLURP1, LYNXI-A et -B sont sécrétées alors que LYPDC2 et LYNX1-C sont liés à la membrane. Trois de ces protéines sont exprimées dans l'épiderme et dans des kératinocytes cultivés. Ces résultats suggèrent que la famille LY6/PLAUR pourrait avoir un rôle important dans l'homéostasie de la peau.

Notas sinonímicas em Lepturini sul-americanos (Coleoptera, Cerambycidae, Lepturinae)

Novas sinonímias propostas: Strangalia flavocincta (Thomson, 1860) = Ophistomis tristis Melzer, 1922 syn. nov. = O. latifasciata Melzer, 1926 syn. nov.; Strangalia succincta (Redtenbacher, 1867) = O. auriflua Redtenbacher, 1867 syn. nov.; Strangalia melanura (Redtenbacher, 1867) = Euryptera dimidiata Redtenbacher, 1867 syn. nov.; Strangalia lyrata (Redtenbacher, 1867) = Ophistomis discophora Redtenbacher, 1867 syn. nov.; Strangalia fulvicornis (Bates, 1872) = Ophistomis variabilis Melzer, 1926 syn. nov. = O. flavovittata Melzer, 1926 syn. nov.; Strangalia melanophthisis (Berg, 1889) reval. = Euryptera melanura var. nigripennis Melzer, 1930 syn. nov.; Anastrangalia sanguinolenta (Linnaeus, 1761) (espécie introduzida na Argentina) = Leptura bonaeriensis Burmeister, 1865 syn. nov.

Insect galls of a protected remnant of the Atlantic Forest tableland from Rio de Janeiro State (Brazil)

ABSTRACT Insect galls of a protected remnant of the Atlantic Forest tableland from Rio de Janeiro State (Brazil): Galling insects in Rio de Janeiro state are known by their great diversity, despite most of the surveys have been done in restinga. This paper investigated the insect galls from a remnant of Atlantic Forest located in São Francisco de Itabapoana municipality, Rio de Janeiro state, Brazil. The galling insect fauna was surveyed from March, 2013 to April, 2014 at the Estação Ecológica Estadual de Guaxindiba. 143 gall morphotypes were found in 31 plant families, 60 genera and 82 species. Fabaceae, Myrtaceae and Sapindaceae were the main host families, being Trichilia, Tontelea and Eugenia the main host genera. Most galls occured on leaves, with globose shape, green and glabrous. Diptera (Cecidomyiidae), Hemiptera, and Lepidoptera were the inducing orders and the associated fauna comprised parasitoids (Hymenoptera), inquilines (Lepidoptera, Coleoptera, and Hemiptera: Coccoidea), successors (Psocoptera, Collembola and Acari), and predators (Pseudoscorpiones). Three plant genera and nine plant species are recorded for the first time as host of galls in Brazil. All the records are new to the municipality, and the distribution of 15 galling species is extended to the North of the state of Rio de Janeiro.

Serie estratigráfica de Liásico de la provincia de Girona

From several small stratigraphic sections, we have reconstituted the Liassic interval of the Girona Province (NE Spain). We have been able to identify paleontologically the Carixian (Jamesoni, Ibex, and perhaps, Davoei zones), the Lower Domerian (Stockesi zone), and the Middle Toarcian (possibily Variabilis zone, as well as Thouarsense and Insigne zones). The Lower Toarcian (Serpentinus zone) is represented by fossil remains no ‘in situ’. The lowest Toarcián (Tenuicos-tatum zone) as well as the Middle and Upper Domerian (Margaritatus and Spinatum zones respectively, are probably represented by stratigraphic gaps

Biodiversity study of arthropods collected on rat carrion in Yaounde, Cameroon: first study on forensic entomology in Central Africa

The first investigation of arthropods associated with carrion in Cameroon was carried out within the campus of the University of Yaounde I (Cameroon) from 17thJanuary to 3rd April 2008. Carcasses of rats (Rattus norvegicus Berkenhout, 1769 var WISTAR) were exposed to colonization by the local fauna of arthropods. The invading organisms were collected daily during the study period. 2287 individuals of arthropod belonging to 3 classes, 16 orders, 37 families and 7 subfamilies were identified. The insects assessed were mainly Diptera, Coleoptera and Acari. This study illustrates the high diversity of the necroentomofauna in Cameroon and provides an insight approximation into the succession pattern of invading insect and a weekly estimation of the time of death.

Effect of brushwood transposition on the leaf litter arthropod fauna in a cerrado area

The results of ecological restoration techniques can be monitored through biological indicators of soil quality such as the leaf litter arthropod fauna. This study aimed to determine the immediate effect of brushwood transposition transferred from an area of native vegetation to a disturbed area, on the leaf litter arthropod fauna in a degraded cerrado area. The arthropod fauna of four areas was compared: a degraded area with signal grass, two experimental brushwood transposition areas, with and without castor oil plants, and an area of native cerrado. In total, 7,660 individuals belonging to 23 taxa were sampled. Acari and Collembola were the most abundant taxa in all studied areas, followed by Coleoptera, Diptera, Hemiptera, Hymenoptera, and Symphyla. The brushwood transposition area without castor oil plants had the lowest abundance and dominance and the highest diversity of all areas, providing evidence of changes in the soil community. Conversely, the results showed that the presence of castor oil plants hampered early succession, negatively affecting ecological restoration in this area.

Sobre el area de dispersion de Artemisia tournefortiana Reich. en la Peninsula Iberica

Al consultar los pliegos de Artemisia L. contenidos en el Herbario BCF, con motivo de la revisión de dicho género en la Península Ibérica, que estamos realizando como tesis doctoral, hemos hallado dos de ellos (BCF n.° 7473 y n.° 7475) recolectados por don Salvador Rivas Goday, el 17 de septiembre de 1948, en Titulcia (MADRID) y determinados como Artemisia variabilis Ten.

Biologia e exigências térmicas do ácaro-vermelho (Tetranychus ludeni Zacher) em folhas de algodoeiro

O objetivo deste trabalho foi estudar a biologia e as exigências térmicas de Tetranychus ludeni Zacher (Acari: Tetranychidae) em folhas de algodoeiro (Gossypium hirsutum L. r. latifolium (Hutch)). Foram utilizadas câmaras climatizadas, ajustadas nas temperaturas de 20, 23, 25, 28 e 30ºC, umidade relativa de 70% e fotófase de 12 horas. Os períodos de desenvolvimento de ovo a adulto variaram de 20,77 (20ºC) a 8,50 dias (30ºC), em fêmeas, e de 18,83 (20ºC) a 7,75 dias (30ºC), em machos. As temperaturasbase, estimadas pelo método da intersecção de X, para os períodos de desenvolvimento de ovoadulto, foram de 14,05ºC para fêmeas e 13,91ºC para machos, enquanto os valores da constante térmica estimados de acordo com a lei de Réamur foram de 138,34 grausdia, para fêmeas e 130,91 grausdia, para machos. Na temperatura de 30ºC foram observados os maiores valores de razão intrínseca de crescimento (0,418), número de ovos de fêmeas/dia (3,47), fecundidade (61,29) e taxa líquida de reprodução (48,00) e o menor valor para o tempo médio de uma geração (9,27).

Toxicidade de extratos de nim (Azadirachta indica) ao ácaro-vermelho-do-cafeeiro Oligonychus ilicis

O objetivo deste trabalho foi estudar a toxicidade aguda e crônica de extratos de óleo de torta, de sementes e de folhas de Azadirachta indica A. Juss (nim) a fêmeas do ácaro-vermelho-do-cafeeiro Oligonychus ilicis (McGregor) (Acari: Tetranychidae). A mortalidade e a taxa instantânea de crescimento populacional de fêmeas adultas desse ácaro foram avaliadas em discos de folhas de cafeeiro com 3,5 cm de diâmetro, impregnados com resíduo seco dos extratos de nim e flutuando em água. As concentrações dos extratos de óleo de torta, sementes e folhas de nim que mataram 50% e 99% dos indivíduos de O. ilicis, após 72 horas de exposição, foram de 0,02, 15,9 e 121,4 mg/mL e de 10,9, 520,9 e 277,4 mg/mL, respectivamente. A taxa instantânea de crescimento populacional de O. ilicis diminuiu, linearmente, com o aumento da concentração dos extratos de óleo de torta, sementes e folhas de nim até 0,075, 15 e 144 mg/mL, respectivamente, a partir das quais as populações desse ácaro foram extintas.

Adubação química, ataque do ácaro Aceria guerreronis e produtividade do coqueiro 'Anão-Verde'

O objetivo deste trabalho foi determinar a influência de níveis de nitrogênio e de potássio aplicados em combinação, via fertirrigação, sobre a produtividade do coqueiro 'Anão-Verde', e a intensidade de ataque do ácaro Aceria guerreronis Keifer (Acari: Eriophyidae). O estudo foi conduzido de fevereiro de 2003 a março de 2004, no Município de Neópolis, SE, em um plantio comercial de coqueiro 'Anão-Verde', com oito anos de idade, que desde o ano 2000 vinha recebendo, via microaspersão, os seguintes tratamentos (gramas de N e K por planta por ano): 135 e 135; 1.890 e 1.890; 810 e 135; 135 e 810; 1.890 e 810; e 810 e 1.890. Foram realizadas quatro avaliações trimestrais, entre abril de 2003 e março de 2004, nos cachos de frutos associados às folhas 14 e 18. A produção de frutos foi influenciada pelos níveis de N e K aplicados ao coqueiral, porém a infestação (8 a 80% de frutos atacados) e a severidade dos danos do ácaro-praga (3 a 47%) não foram afetadas pela adubação química.

Occurrence of Aceria tosichella in Brazil

The objective of this work was to evaluate the occurrence of Aceria tosichella Keifer (Prostigmata: Eriophyidae), the wheat curl mite, in Rio Grande do Sul, Brazil. Samples to detect A. tosichella specimens were collected in wheat, corn, oat crops and potential host grasses in 46 localities, in October 2006, August and October 2007. Samples of wheat were also collected in experimental greenhouses at Embrapa Trigo, Passo Fundo, RS, Brazil. A. tosichella specimens were found in wheat samples from Passo Fundo, Palmeira das Missões, São Luís Gonzaga, and Santo Antônio das Missões, RS, Brazil. Symptoms due to A. tosichella infestations were observed only in greenhouse conditions. This is the first report of A. tosichella in Brazil and the second in South America.

Predação por coccinelídeos e crisopídeo influenciada pela teia de Tetranychus evansi

O objetivo deste trabalho foi avaliar o efeito da teia produzida por Tetranychus evansi (Acari: Tetranychidae) na eficiência de predação por coccinelídeos Cycloneda sanguinea e Eriopis connexa e pelo crisopídeo Chrysoperla externa. A eficiência em predar as fêmeas de T. evansi na presença e na ausência de sua teia em discos de folhas de tomateiro com ácaros foi avaliada para cada instar das diferentes espécies de predadores. Foi observado efeito negativo da teia produzida por T. evansi na eficiência de predação do segundo e do terceiro instar de C. externa. Quanto à E. connexa, somente a predação pelo último instar foi influenciada negativamente pela presença da teia. A predação pelas larvas de primeiro e terceiro instar de C. sanguinea foi significativamente menor na presença da teia; no entanto, o resultado foi contrário para o quarto instar. O efeito da teia de T. evansi como mecanismo de defesa varia conforme a espécie e o estágio de desenvolvimento do predador.

Comparação econômica entre controle biológico e químico para o manejo de ácaro-vermelho em macieira

O ácaro-vermelho da macieira, Panonychus ulmi (Acari: Tetranychidae), é uma importante praga na cultura da macieira em Fraiburgo - SC, e o controle biológico aplicado foi implantado em meados dos anos 90. O objetivo deste trabalho foi demonstrar os benefícios econômicos da utilização do controle biológico no manejo do ácaro-vermelho. A avaliação foi realizada em dois pomares comerciais de macieiras. Em um deles, foi implantado o controle biológico aplicado de ácaros, baseado na liberação do ácaro predador Neoseiulus californicus (Acari: Phytoseiidae), seleção de inseticidas e manejo de ervas invasoras, e o outro pomar seguiu o manejo convencional de artrópodes, baseado na aplicação de produtos químicos para o controle de insetos, ácaros fitófagos e ervas invasoras. A análise econômica mostrou que os custos com mão-de-obra e máquinas foram semelhantes em ambos os pomares, entretanto os custos com acaricidas foram significativamente inferiores no pomar onde o manejo foi o controle biológico, demonstrando que, apesar da necessidade de investimentos em instalações para a criação do ácaro predador e custos de manutenção das mesmas, a estratégia biológica foi economicamente viável.

Mites fluctuation population on peach tree (Prunus persica (L.) Batsch) and in associated plants

Despite the importance of peach (Prunus persica (L.) Batsch) in Rio Grande do Sul, little is known about mites fluctuation population considered important to this crop. The objective of this study was to know the population diversity and fluctuation of mite species associated with Premier and Eldorado varieties in Roca Sales and Venâncio Aires counties, Rio Grande do Sul. The study was conducted from July 2008 to June 2009 when 15 plants were randomly chosen in each area. The plants were divided in quadrants and from each one a branch was chosen from which three leaves were removed: one collected in the apical region, another in the medium and the other in the basal region, totalizing 180 leaves/area. Five of the most abundant associated plants were collected monthly in enough amounts for the screening under the stereoscopic microscope during an hour. A total of 1,124 mites were found belonging to 14 families and 28 species. Tetranychus ludeni Zacher, 1913, Panonychus ulmi (Koch, 1836) and Mononychellus planki (McGregor, 1950) were the most abundant phytophagous mites, whereas Typhlodromalus aripo Deleon, 1967 and Phytoseiulus macropilis (Banks, 1904) the most common predatory mites. The period of one hour under stereoscopic microscope was enough to get a representative sample. In both places evaluated the ecologic indices were low, but little higherin Premier (H' 0.56; EqJ: 0.43) when compared to Eldorado (H' 0.53; EqJ 0.40). In Premier constant species were not observed and accessory only Brevipalpus phoenicis (Geijskes, 1939), T. ludeni and T. aripo. Higher abundance was observed in December and January and bigger amount in April. Already in Eldorado, T. ludeni and P. ulmi were constants. Greater abundance was observed in November and December, whereas grater richness in December and January. In both orchards were not found mites in buds. Tetranychus ludeni is the most abundant phytophagous mites with outbreak population in November, December and January and high predator diversity was observed on associated plants and on peach plants, indicating the existence of species mobility in peach orchard.

CARACTERIZAÇÃO QUÍMICA E EFEITO DO ÓLEO ESSENCIAL DE ERVA-DE-SANTA-MARIA SOBRE O ÁCARO-RAJADO DE MORANGUEIRO

O ácaro-rajado, Tetranychus urticae Koch (1836) (Acari: Tetranychidae), é a principal praga na cultura do morangueiro. O objetivo deste trabalho foi a caracterização química do óleo essencial de Chenopodium ambrosioides L. e a avaliação de seus efeitos sobre o ácaro-rajado. Os compostos do óleo essencial foram identificados pelasanálises de CG-DIC e CG-EM. A mortalidade e a oviposição foram avaliadas após a pulverização do óleo (2,5; 5,0; 7,5 e 10,0% v v-1) sobre fêmeas do ácaro-rajado. O rendimento do óleo essencial foi de 0,3%, e os compostos identificados foram (Z)-ascaridol (87%), (E)-ascaridol (5,04%), p-cimeno (4,83%), a- terpineno (1,24%) e piperitona (0,7%). Os maiores valores para mortalidade e redução de oviposição observados foram 85% e 69%, respectivamente. O óleo essencial de C. ambrosioidesé uma opção promissora para o manejo do ácaro-rajado.