Characterisation of an IS1 induced mutation in the carboxy-terminal end of bacteriophage Mu transposase which affects several functional domains of the protein

We show that a mutation in bacteriophage Mu transposase (pA) which was isolated as a deletion of the C-terminal end of the protein actually consists of the replacement of the last 16 amino acids (which are mostly hydrophilic) by 26 mostly hydrophobic amino acids. This change almost completely inactivates the in vivo enzyme activity as well as its capacity to bind Mu ends in vitro, although the end-binding domain of the protein resides at least 150 amino acids from the C-terminus. This sharply contrasts with the properties of a series of missense mutations and short C-terminal deletions in pA described earlier which only slightly decrease the overall transposase activity.     

Bax-inhibiting peptide protects cells from polyglutamine toxicity caused by Ku70 acetylation

Polyglutamine (polyQ) diseases, such as Huntington's disease and Machado-Joseph disease (MJD), are caused by gain of toxic function of abnormally expanded polyQ tracts. Here, we show that expanded polyQ of ataxin-3 (Q79C), a gene that causes MJD, stimulates Ku70 acetylation, which in turn dissociates the proapoptotic protein Bax from Ku70, thereby promoting Bax activation and subsequent cell death. The Q79C-induced cell death was significantly blocked by Ku70 or Bax-inhibiting peptides (BIPs) designed from Ku70. Furthermore, expression of SIRT1 deacetylase and the addition of a SIRT1 agonist, resveratrol, reduced Q79C toxicity. In contrast, mimicking acetylation of Ku70 abolished the ability of Ku70 to suppress Q79C toxicity. These results indicate that Bax and Ku70 acetylation play important roles in Q79C-induced cell death, and that BIP may be useful in the development of therapeutics for polyQ diseases.     

Evaluation of the sticky MosquiTRAP for detecting Aedes (Stegomyia) aegypti (L.) (Diptera: Culicidae) during the dry season in Belo Horizonte, Minas Gerais, Brazil

MosquiTRAP is a sticky trap specifically designed to capture gravid females of Aedes aegypti (L.) and allows the identification of the mosquito in the field during the inspection of the trap. This study aims to compare this sticky trap to larval and ovitrap surveys for field monitoring of A. aegypti during the dry season. The study was conducted from March to June of 2003 in 20 blocks of the district of Itapo?, Belo Horizonte, MG. The traps were monitored every week while the larval survey was conducted on a monthly basis. The larval index: Premise Index (PI) and Breteau Index (BI) had equal values throughout the experiment (1.72 in the first two months and zero in the last two). The container index (CI) during the first two months was 0.09 and 0.1%, respectively and zero in the last two. The Ovitrap Positive Index (OPI) ranged from 16.7 to 76.9%, and the MosquiTRAP Positive Index (MPI) ranged from 0 to 31.5%. The Egg Density Index (EDI) ranged from 26.6 to 82.8, while the Adult Density Index ranged from 0 to 1.6 throughout the experiment. Temperature and rainfall did not affect the Positive and Density Indices, although these environmental variables seemed to have affected the larvae indices. Although the MosquiTRAP caught a low number of Aedes mosquitoes during the study, it was more sensitive than the larval survey to detect the presence of Aedes mosquitoes.     

Marked change in the balance between CYP27A1 and CYP46A1 mediated elimination of cholesterol during differentiation of human neuronal cells

Cholesterol metabolism in the brain is distinct from that in other tissues due to the fact that cholesterol itself is unable to pass across the blood-brain barrier. Elimination of brain cholesterol is mainly dependent on a neuronal-specific cytochrome P450, CYP46A1, catalyzing the conversion of cholesterol into 24(S)-hydroxycholesterol (24OHC), which is able to pass the blood-brain barrier. A suitable model for studying this elimination from human neuronal cells has not been described previously. It is shown here that differentiated Ntera2/clone D1 (NT2) cells express the key genes involved in brain cholesterol homeostasis including CYP46A1, and that the expression profiles of the genes observed during neuronal differentiation are those expected to occur in vivo. Thus there was a decrease in the mRNA levels corresponding to cholesterol synthesis enzymes and a marked increase in the mRNA level of CYP46A1. The latter increase was associated with increased levels of CYP46A1 protein and increased production of 24OHC. The magnitude of the secretion of 24OHC from the differentiated NT2 cells into the medium was similar to that expected to occur under in vivo conditions. An alternative to elimination of cholesterol by the CYP46A1 mechanism is elimination by CYP27A1, and the product of this enzyme, 27-hydroxycholesterol (27OHC), is also known to pass the blood-brain barrier. The CYP27A1 protein level decreased during the differentiation of the NT2 cells in parallel with decreased production of 27OHC. The ratio between 24OHC and 27OHC in the medium from the cultured cells increased, by a factor of 13, during the differentiation process. The results suggest that progenitor cells eliminate cholesterol in the form of 27OHC while neurogenesis induces a change to the CYP46A1 dependent pathway. Furthermore this study demonstrates that differentiated NT2 cells are suitable for studies of cholesterol homeostasis in human neurons.     

Genetic characterization of Latin-American Creole cattle using microsatellite markers

Genetic diversity in and relationships among 26 Creole cattle breeds from 10 American countries were assessed using 19 microsatellites. Heterozygosities, F-statistics estimates, genetic distances, multivariate analyses and assignment tests were performed. The levels of within-breed diversity detected in Creole cattle were considerable and higher than those previously reported for European breeds, but similar to those found in other Latin American breeds. Differences among breeds accounted for 8.4% of the total genetic variability. Most breeds clustered separately when the number of pre-defined populations was 21 (the most probable K value), with the exception of some closely related breeds that shared the same cluster and others that were admixed. Despite the high genetic diversity detected, significant inbreeding was also observed within some breeds, and heterozygote excess was detected in others. These results indicate that Creoles represent important reservoirs of cattle genetic diversity and that appropriate conservation measures should be implemented for these native breeds in order to minimize inbreeding and uncontrolled crossbreeding.     

Spatial Patterns of Benthic Macroinvertebrates in Intertidal Areas of a Southern European Estuary: The Tagus, Portugal

This study characterizes the composition and spatial distribution patterns of the benthic macrofauna in the intertidal mudflats of the Tagus estuary, western Portugal. A total of 68 species, more than 226,000 specimens with a total wet weight biomass of approximately 1170 g were identified in 380 sites. The species Streblospio shrubsolii, Cyathura carinata, Tharyx sp., Hydrobia ulvae and Tubificids were the most common and abundant. Scrobicularia plana strongly dominated the biomass. The invertebrate macrofauna of the Tagus estuary shows similarities to what is known from other temperate mudflats. The diversity of species, their overall abundance and the ratio of Molluscs plus Crustaceans to Polychaete species corroborate the distinctiveness between temperate and tropical mudflats and sandflats. The spatial distribution of the fauna reflects the sediment characteristics but the relationship between the environmental and the biological data is not as strong as obtained for sublittoral areas. This relationship diminishes from the sublittoral shelf to sublittoral estuarine areas, showing minimum values in this study, suggesting that such a relationship is less straightforward as natural disturbance increases. Nevertheless, a mixture of grain-size, elevation (inundation time) and particular habitats (relic oyster beds) form the best explanatory factors for the spatial distribution patterns of the intertidal benthic macrofauna of the Tagus estuary.     

Genetic variation in BoLA microsatellite loci in Portuguese cattle breeds

Major histocompatibility complex (MHC) typing based on microsatellites can be a valuable approach to understanding the selective processes occurring at linked or physically close MHC genes and can provide important information on variability and relationships of populations. Using microsatellites within or in close proximity with bovine lymphocyte antigen (BoLA) genes, we investigated the polymorphisms in the bovine MHC, known as the BoLA, in eight Portuguese cattle breeds. Additional data from non-BoLA microsatellite loci were also used to compare the variability between these regions. Diversity was higher in BoLA than in non-BoLA microsatellites, as could be observed by the number of alleles, allelic richness and observed heterozygosity. Brava de Lide, a breed selected for aggressiveness and nobility, presented the lowest values of observed heterozygosity and allelic richness in both markers. Results from neutrality tests showed few statistically significant differences between the observed Hardy–Weinberg homozygosity ( F ) and the expected homozygosity ( F _E), indicating the apparent neutrality of the BoLA microsatellites within the analysed breeds. Nevertheless, we detected a trend of lower values of observed homozygosity compared with the expected one. We also detected some differences in the levels of allelic variability among the four BoLA microsatellites. Our data showed a higher number of alleles at the BoLA-DRB3 locus than at the BoLA-DRBP1 locus. These differences could be related to their physical position in the chromosome and may reflect functional requirements for diversity.     

Diversity of filamentous fungi associated with Hypothenemus hampei (Ferrari) (Coleoptera: Scolytidae) and its galleries in berries of Coffea canephora (Pierre)

Field sampling was carried out in Ouro Preto d'Oeste - Rond?nia (10 degrees 45'S and 62 degrees 15'W) to evaluate the mycobiota associated with Hypothenemus hampei Ferrari [cuticle, mouth, prothorax (mycangia), gut and feces] and its galleries on berries of Coffea canephora Pierre. Ten genera (201 isolates) were directly related with the insect while five genera (20 isolates) were related with galleries on berries. All the genera identified in the insects were also present in their galleries, what indicates that boring may be an active way of fungi inoculation by H. hampei. The fungi genera were more diverse in the mouth and prothorax of borers, and lower in feces. Fusarium, Penicillium and Geotrichum, with abundance of 55.7, 24.3 and 10.8%, respectively, were dominant genera. In the galleries Fusarium, Geotrichum, Trichoderma and Aspergillus with abundance of 33.3, 29.6, 18.5 and 14.8%, respectively, were dominant genera. The overall presence of Fusarium in coffee berry borer and its galleries) reinforces previous indications of a close interaction between H. hampei-Fusarium. The presence of Aspergillus and Penicillium emphasizes the possibility of "ochratoxin dispersion" by the borer. This work provides the first record of the mycobiota associated with H. hampei in C. canephora. Among the identified genera, Cephalosporium, Geotrichum and Oidiodendrum were recorded for the first time in association with H. hampei and its galleries in C. canephora.