In vivo binding of the Cry11Bb toxin of Bacillus thuringiensis subsp. medellin to the midgut of mosquito larvae (Diptera: Culicidae)

Bacillus thuringiensis subsp. medellin produces numerous proteins among which 94 kDa known as Cry11Bb, has mosquitocidal activity. The mode of action of the Cry11 proteins has been described as similar to those of the Cry1 toxins, nevertheless, the mechanism of action is still not clear. In this study we investigated the in vivo binding of the Cry11Bb toxin to the midgut of the insect species Anopheles albimanus, Aedes aegypti, and Culex quinquefasciatus by immunohistochemical analysis. Spodoptera frugiperda was included as negative control. The Cry11Bb protein was detected on the apical microvilli of the midgut epithelial cells, mostly on the posterior midgut and gastric caeca of the three mosquito species. Additionally, the toxin was detected in the Malpighian tubules of An. albimanus, Ae. aegypti, Cx. quinquefasciatus, and in the basal membrane of the epithelial cells of Ae. aegypti midgut. No toxin accumulation was observed in the peritrophic membrane of any of the mosquito species studied. These results confirm that the primary site of action of the Cry11 toxins is the apical membrane of the midgut epithelial cells of mosquito larvae.

Histopathological and ultrastructural effects of delta-endotoxins of Bacillus thuringiensis serovar israelensis in the midgut of Simulium pertinax larvae (Diptera, Simuliidae)

The bacterium Bacillus thuringiensis (Bt) produces parasporal crystals containing delta-endotoxins responsible for selective insecticidal activity on larvae. Upon ingestion, these crystals are solubilized in the midgut lumen and converted into active toxins that bind to receptors present on the microvilli causing serious damage to the epithelial columnar cells. We investigated the effect of these endotoxins on larvae of the Simulium pertinax, a common black fly in Brazil, using several concentrations during 4 h of the serovar israelensis strain IPS-82 (LFB-FIOCRUZ 584), serotype H-14 type strain of the Institute Pasteur, Paris. Light and electron microscope observations revealed, by time and endotoxin concentration, increasing damages of the larvae midgut epithelium. The most characteristic effects were midgut columnar cell vacuolization, microvilli damages, epithelium cell contents passing into the midgut lumen and finally the cell death. This article is the first report of the histopathological effects of the Bti endotoxins in the midgut of S. pertinax larvae and the data obtained may contribute to a better understanding of the mode of action of this bacterial strain used as bioinsecticide against black fly larvae.

Comparison of two commercial formulations of Bacillus thuringiensis var. israelensis for the control of Anopheles aquasalis (Diptera: Culicidae) at three salt concentrations

Anopheles aquasalis larvae are salt water tolerant, preferring concentrations between 10 and 20 parts per thousand (ppt). The larvicidal efficacy of two formulations of Bacillus thuringiensis var. israelensis (Vectobac-12AS® and Bactivec®), was investigated against An. aquasalis at salinities of 0, 10, and 20 ppt. A probit analysis was used to calculate the lethal concentrations (LC50 and LC95) for each product at each salinity. The LC50 and LC95 were higher for Bactivec® than Vectobac-12AS®, and for Bactivec®, the LC50 and LC95 increased with salinity. Vectobac-12AS® should thus be preferred to Bactivec® for An. aquasalis control, especially in saline breeding habitats.

Tracing lineage by phenotypic and genotypic markers in Salmonella enterica subsp. enterica serovar 1,4,[5],12:i:- and Salmonella Typhimurium isolated in state of São Paulo, Brazil

Fifty-three Salmonella 1,4,[5],12:i:- and 45 Salmonella Typhimurium strains were characterised using phage typing, plasmid profiles and pulsed-field gel electrophoresis (PFGE) for comparison. The majority of the strains were subdivided into definitive type (DT) 41 (22.6%) and DT 193 (18%) and the 60-MDa plasmid was detected in 94.3% and 84.4% of strains, respectively. Genetic diversity was observed among all strains and 90% presented a > 70% similarity through PFGE analysis. These results suggest a close relationship between Salmonella 1,4,[5],12:i:- and Salmonella Typhimurium at the serotype level.

Rapid tests for the detection of the Mycobacterium abscessus subsp. bolletii strain responsible for an epidemic of surgical-site infections in Brazil

A single strain of Mycobacterium abscessus subsp. bolletii, characterised by a particular rpoB sequevar and two highly related pulsed field gel electrophoresis patterns has been responsible for a nationwide outbreak of surgical infections in Brazil since 2004. In this study, we developed molecular tests based on polymerase chain reaction restriction-enzyme analysis (PRA) and sequencing for the rapid identification of this strain. Sequences of 15 DNA regions conserved in mycobacteria were retrieved from GenBank or sequenced and analysed in silico. Single nucleotide polymorphisms specific to the epidemic strain and located in enzyme recognition sites were detected in rpoB, the 3' region of the 16S rDNA and gyrB. The three tests that were developed, i.e., PRA-rpoB, PRA-16S and gyrB sequence analysis, showed 100%, 100% and 92.31% sensitivity and 93.06%, 90.28% and 100% specificity, respectively, for the discrimination of the surgical strain from other M. abscessus subsp. bolletii isolates, including 116 isolates from 95 patients, one environmental isolate and two type strains. The results of the three tests were stable, as shown by results obtained for different isolates from the same patient. In conclusion, due to the clinical and epidemiological importance of this strain, these tests could be implemented in reference laboratories for the rapid preliminary diagnosis and epidemiological surveillance of this epidemic strain.

Polygonum Romanum jacq. subsp. Gallicum (Raffaelli) Raffaelli & villar Polygonaceae) en el Ampurdán (Gerona)

Article que descriu la planta poligonàcea, Polyganum Romanum, present a la zona de l' Empordà

TELINE STENOPETALA (WEBB & BERTHEL.) WEBB & BERTHEL. SUBSP. SANTOANTAOI SUBSP. NOV., PARA SANTO ANTÃO, ISLAS DE CABO VERDE

Se describe una nueva subespecie de Teline stenopetala (Webb & Berthel.) Webb & Berthel.: T. stenopetala subsp. santoantaoi Marrero-Rodr. subsp. nov., para la isla de Santo Antão, Islas de Cabo Verde. Se comenta las relaciones taxonómicas con las otras subespecies descritas. Igualmente se hace un breve comentario sobre su hábitat, ecología y estado de conservación de sus poblaciones.

Estudo Químico e Biológico das Espécies Vegetais Caboverdianas Echium Hypertropicum Webb e Echium Stenosiphon Webb Subsp. Stenosiphon

Echium hypertropicum Webb e Echium stenosiphon Webb subsp. stenosiphon são arbustos endêmicos de Cabo Verde, usados na medicina popular para o tratamento de distúrbios gastrintestinais e tosse. As duas espécies tiveram suas frações alcalóidicas obtidas por extração ácido-base. A análise por CG-EM e ESI-EM/EM indicou a presença de alcaloides pirrolizidínicos (APs) e as substâncias purificadas foram analizadas por experimentos de RMN de 1D e 2D. Um total de 10 alcaloides foram isoladas e identificadas, sendo que 8 identificadas através da comparação de suas massas moleculares e padrões de fragmentação de massas, com a base de dados NIST e os dados da litratura para o género. Os diésteres hepatotóxicos equimidana e 7-(2-metilbutiril)-9-equimidinilretronecina foram identificadas em ambas as espécies. Os alcaloides 7-senecioilretronecina, 9-angeloilretronecina, licopsamina, 7-acetil-licopsamina e equihumilina foram identificados nas folhas de E. hypertropicum, enquanto que o N-óxido da 7-(2-metilbutiril)-9-equimidinilretronecina foi identificado nas folhas de E. stenosipnhon. A equimidina foi o componente majoritário na fração em éter dietílico das folhas de E. hypertropicum, enquanto a 7-(2-metilbutiril)-9-equimidinilretronecina foi o componente majoritário na fração em diclorometano das folhas de E. stenosiphon. O alcaloide 7-(2-metilbutiril)-9-equimidinilretronecina N-óxido foi identificado pela primeira vez no gênero Echium. Em adição, 22 componentes de óleo essencial foram identificadas nas flores de Echium hypertropicum, sendo trans-fitol (30,64 %), n-pentacosano (8,28 %) e n-tricosano (6,73) como componentes majoritários. O triterpeno friedelina foi também isolado das folhas de E. hypertropicum. Na avaliação da atividade antibacteriana, os extratos etanólicos das duas espécies vegetais e o alcaloide 7-(2-metilbutiril)-9-equimidinilretronecina foram capazes de inibir o crescimento de Staphylococcus aureus ATCC 29213 com CMI de 250,0 μg/mL e 25,0 μg/mL, respectivamente. A atividade anticolinesterásica foi avaliada e a equimidina foi capaz de inibir a enzima acetilcolinesterase nas concentrações testadas com o valor de P = 0,0011. O alcaloide 7-(2-metilbutiril)-9-equimidinilretronecina retardou o crescimento do fitófago Dysdercus peruvianus na concentração de 1mg/mL. Os extratos etanólicos de E. hypertropicum e E. stenosiphon (3,9 μg/mL) foram avaliados frente ao vírus HSV. O extrato etanólico de E. hypertropicum apresentou uma porcentagem de inibição (PI) de 27,5% contra HSV-1S e 43,8% contra HSV-2S. Apresentaram ainda elevada citotoxidade para as celulas Vero, utilizadas como sistema hospedeiro (CC50 de 140,10 μg/mL e 96,86 μg/mL). A composição química e as atividades biológicas de E. hypertropicum e E. stenosiphon subsp. stenosiphon foram relatadas pela primeira vez. As substâncias identificadas podem ser utilizadas no futuro como marcadores quimiotaxonômicos para o gênero Echium.

Dynamics of the control of Aedes (Stegomyia) aegypti Linnaeus (Diptera, Culicidae) by Bacillus thuringiensis var israelensis, related with temperature, density and concentration of insecticide

The dynamics of the control of Aedes (Stegomyia) aegypti Linnaeus, (Diptera, Culicidae) by Bacillus thuringiensis var israelensis has been related with the temperature, density and concentration of the insecticide. A mathematical model for biological control of Aedes aegypti with Bacillus thuringiensis var israelensis (Bti) was constructed by using data from the literature regarding the biology of the vector. The life cycle was described by differential equations. Lethal concentrations (LC50 and LC95) of Bti were determined in the laboratory under different experimental conditions. Temperature, colony, larvae density and bioinsecticide concentration presented marked differences in the analysis of the whole set of variables; although when analyzed individually, only the temperature and concentration showed changes. The simulations indicated an inverse relationship between temperature and mosquito population, nonetheless, faster growth of populations is reached at higher temperatures. As conclusion, the model suggests the use of integrated control strategies for immature and adult mosquitoes in order to achieve a reduction of Aedes aegypti.

Isolamento de esporos de Paenibacillus larvae subsp. larvae no Brasil

Este trabalho objetivou detectar presença de esporos de Paenibacillus larvae subsp. larvae em produtos de um entreposto do interior do Estado do Rio Grande do Sul, a identificação de possíveis fontes de contaminação e a avaliação da possibilidade da transferência de esporos para colméias de apiários adjacentes a partir de produtos importados contaminados. Foram analisados mel e pólen importados disponíveis no entreposto, favo do ninho (crias, pólen e mel) colhido de uma colméia sadia, mel estocado em um dos apiários e abelhas adultas. Os resultados foram positivosem relação ao mel e pólen importados, a três grupos de abelhas adultas e ao mel do favo.

Estudi de la població de Spiraea crenata subsp. parvifolia (Rosaceae) del Collsacabra

Spiraea crenata subsp. parvifolia és un tàxon endèmic del nord-est ibèric, que té una de les escasses poblacions conegudes al Collsacabra, Osona. En aquest treball es pretén definir l’extensió d’aquesta població, caracteritzar la seva estructura demogràfica i avaluar-ne l’estat de conservació. La majoria d’individus censats viuen en un nucli principal rodejat per nuclis més petits. L’estabilitat de l’ambient que ocupa permet valorar l’evolució de la població i la categoria UICN d’amenaça, actualment considerada «en perill ». Es proposen mesures de gestió que permetin la conservació d’aquesta població.

Mycobacterium avium subsp. paratuberculosis in an Italian Cohort of Type 1 Diabetes Pediatric Patients.

Mycobacterium avium subsp. paratuberculosis (MAP) is the etiological agent of Johne's disease in ruminants. Recent studies have linked MAP to type 1 diabetes (T1D) in the Sardinian population. The aim of this study was to investigate the prevalence of MAP infection in a T1D cohort from continental Italy compared with healthy control subjects. 247 T1D subjects and 110 healthy controls were tested for the presence of MAP. MAP DNA was detected using IS900-specific polymerase chain reaction (PCR). The presence of antibodies towards a MAP antigen, heparin binding hemoagglutinin (HBHA), was detected by ELISA. We demonstrated a higher MAP DNA prevalence in plasma samples from T1D patients and a stronger immune response towards MAP HBHA, compared with healthy control subjects. Moreover, in the recent onset patients, we observed an association between anti-MAP antibodies and HLA DQ2 (DQA1 0201/DQB1 0202). These findings taken together support the hypothesis of MAP as an environmental risk factor for the development of T1D in genetically predisposed subjects, probably involving a mechanism of molecular mimicry between MAP antigens and pancreatic islet β-cells.

Redução dos sintomas causados pela Xylella fastidiosa subsp. pauca por meio de aplicação de benzotiadiazole e silício

O objetivo deste trabalho foi avaliar o efeito da aplicação do benzotiadiazole (BTH) e do silício sobre o controle da doença causada pela Xylella fastidiosa subsp. pauca em Nicotiana tabacum. Os experimentos foram conduzidos em condições de casa de vegetação, onde as plantas de N. tabacum receberam inoculação de X. fastidiosa (linhagem 9a5c) com 4,7x10(7) UFC mL-1. Os tratamentos consistiram da pulverização das folhas, com soluções de BTH (0,6 e 1,2 mM), e aplicação ao solo de soluções de metassilicato de sódio (2 e 4 µM de Si). Cinco plantas foram utilizadas, por tratamento. Plantas de N. tabacum tratadas com BTH não demonstraram redução de sintomas da bacteriose. Entretanto, plantas tratadas com metassilicato de sódio, sim. A indução de resistência pelo Si poderá ser útil no controle da clorose variegada dos citros.

Detection of siderophores in endophytic bacteria Methylobacterium spp. associated with Xylella fastidiosa subsp. pauca

The objective of this work was to study the production of siderophores by endophytic bacteria Methylobacterium spp., which occupy the same ecological niche as Xylella fastidiosa subsp. pauca (Xfp) in citrus plants. The siderophore production of Methylobacterium strains was tested according to chromeazurol agar assay test (CAS), Csáky test (hydroxamate-type) and Arnow test (catechol-type). In addition, the ability of Xfp to use siderophores, in vitro, produced by endophytic bacteria as source of iron, was evaluated. All 37 strains of Methylobacterium spp. tested were CAS-positive for siderophore production. Methylobacterium spp. produced hydroxamate-type, but not catechol-type siderophores. In vitro growth of Xfp was stimulated by the presence of supernatant siderophores of endophytic Methylobacterium mesophilicum.