Localization of sequences regulating ancestral and acquired sites of esterase6 activity in Drosophila melanogaster

We have broadly defined the DNA regions regulating esterase6 activity in several life stages and tissue types of D. melanogaster using P- element-mediated transformation of constructs that contain the esterase6 coding region and deletions or substitutions in 5' or 3' flanking DNA. Hemolymph is a conserved ancestral site of EST6 activity in Drosophila and the primary sequences regulating its activity lie between -171 and -25 bp relative to the translation initiation site: deletion of these sequences decrease activity approximately 20-fold. Hemolymph activity is also modulated by four other DNA regions, three of which lie 5' and one of which lies 3' of the coding region. Of these, two have positive and two have negative effects, each of approximately twofold. Esterase6 activity is present also in two male reproductive tract tissues; the ejaculatory bulb, which is another ancestral activity site, and the ejaculatory duct, which is a recently acquired site within the melanogaster species subgroup. Activities in these tissues are at least in part independently regulated: activity in the ejaculatory bulb is conferred by sequences between -273 and -172 bp (threefold decrease when deleted), while activity in the ejaculatory duct is conferred by more distal sequences between -844 and -614 bp (fourfold decrease when deleted). The reproductive tract activity is further modulated by two additional DNA regions, one in 5' DNA (-613 to -284 bp; threefold decrease when deleted) and the other in 3' DNA (+1860 to +2731 bp; threefold decrease when deleted) that probably overlaps the adjacent esteraseP gene. Collating these data with previous studies suggests that expression of EST6 in the ancestral sites is mainly regulated by conserved proximal sequences while more variable distal sequences regulate expression in the acquired ejaculatory duct site.      

Nucleotide variation at the hypervariable esterase 6 isozyme locus of Drosophila simulans

Esterase 6 (Est-6/EST6) is polymorphic in both Drosophila melanogaster and D. simulans for two common allozyme forms, as well as for several other less common variants. Parallel latitudinal clines in the frequencies of the common EST6-F and EST6-S allozymes in these species have previously been interpreted in terms of a shared amino acid polymorphism that distinguishes the two variants and is subject to selection. Here we compare the sequences of four D. simulans Est-6 isolates and show that overall estimates of nucleotide heterozygosity in both coding and 5' flanking regions are more than threefold higher than those obtained previously for this gene in D. melanogaster. Nevertheless, the ratio of replacement to exon silent-site polymorphism in D. simulans is less than the ratio of replacement to silent divergence between D. simulans and D. melanogaster, which could be the result of increased efficiency of selection against replacement polymorphisms in D. simulans or to divergent selection between the two species. We also find that the amino acid polymorphisms separating EST6- F and EST6-S in D. simulans are not the same as those that separate these allozymes in D. melanogaster, implying that the shared clines do not reflect shared molecular targets for selection. All comparisons within and between the two species reveal a remarkable paucity of variation in a stretch of nearly 400 bp immediately 5' of the gene, indicative of strong selective constraint to retain essential aspects of Est-6 promoter function.      

The nucleotide sequence of Euglena cytoplasmic phenylalanine transfer RNA. Evidence for possible classifications of Euglena among the animal rather than the plant kingdom

The nucleotide sequence of cytoplasmic phenylalanine tRNA from Euglena gracilis has been elucidated using procedures described previously for the corresponding chloroplastic tRNA [Cell, 9, 717 (1976)]. The sequence is: pG-C-C-G-A-C-U-U-A-m(2)G-C-U-Cm-A-G-D-D-G-G-G-A-G-A-G-C-m(2)2G-psi-psi-A-G-A-Cm -U-Gm-A-A-Y-A-psi-C-U-A-A-A-G-m(7)G-U-C-*C-C-U-G-G-T-psi-C-G-m(1)A-U-C-C-C-G-G- G-A-G-psi-C-G-G-C-A-C-C-A. Like other tRNA Phes thus far sequenced, this tRNA has a chain length of 76 nucleotides. The sequence of E. gracilis cytoplasmic tRNA Phe is quite different (27 nucleotides out of 76 different) from that of the corresponding chloroplastic tRNA but is surprisingly similar (72 out of 76 nucleotides identical) to that of tRNA Phe from mammalian cytoplasm. This extent of sequence homology even exceeds that found between E. gracilis and wheat germ cytoplasmic tRNA Phe. These findings raise interesting questions on the evolution of tRNAs and the taxonomy of Euglena.     

Antimicrobial and antiviral activity-guided fractionation from Scutia buxifolia Reissek extracts

Antimicrobial and antiviral activities of the fractions from Scutia buxifolia stem bark and leaves were evaluated. Best antimicrobial results occurred with the ethyl acetate (EA) and n-butanolic (NB) fractions from the leaves against Micrococcus sp. (minimal inhibitory concentration—MIC = 62.5 μg/ml), and NB fraction from stem bark and leaves against Klebsiella pneumoniae and Enterococcus faecalis (MIC = 62.5 μg/ml). The most active fractions were selected and fractioned into silica column to perform an in vitro antibiofilm assay, which evidenced subfractions EA2 and EA3 as the more active against Candida albicans (biofilm inhibitory concentration—BIC = 582 ± 0.01 μg/ml) and Staphylococcus aureus (BIC = 360 ± 0.007 μg/ml), respectively. The NB (selectivity index—SI = 25.78) and the EA (SI = 15.97) fractions from the stem bark, and the EA (SI = 14.13) fraction from the leaves exhibited a potential antiviral activity towards Herpes Simplex Virus type 1 whereas EA2 and EA3 subfractions from leaves (SI = 12.59 and 10.06, respectively), and NB2 subfraction from stem bark (SI = 12.34) maintained this good activity. Phenolic acids and flavonoids (gallic acid, chlorogenic acid, caffeic acid, rutin, isoquercitrin, quercitrin and quercetin) were identified by HPLC and may be partially responsible for the antimicrobial and antiherpes activities observed. The results obtained in this study showed that Scutia buxifolia has antibiofilm and anti-herpetic activities and that these properties are reported for the first time for this species.     

Defaunation and fragmentation erode small mammal diversity dimensions in tropical forests

Forest fragmentation and defaunation are considered the main drivers of biodiversity loss, yet the synergistic effects of landscape changes and biotic interactions on assemblage structure have been poorly investigated. Here, we use an extensive dataset of 283 assemblages and 105 species of small mammals to understand how defaunation of medium and large mammals and forest fragmentation change the community composition and diversity of rodents and marsupials in tropical forests of South America. We used structured equation models to investigate the relationship between small mammal species, functional and phylogenetic diversity with forest size, forest cover and the occurrence of medium and large mammals. The best‐fit model showed that defaunation reduced functional diversity, and that species diversity of small mammals increased with forest patch size. Forest cover did not affect functional and phylogenetic diversity. Our results indicate that occurrence of medium and large sized mammals (probably acting as predators, or competitors of small mammals) and forest patch size help to retain species and functional diversity in small mammal communities. Further, the number of species in a small mammal community was critical to the maintenance of phylogenetic diversity, and may have a pronounced influence on the ecological functions played by small mammals. Identifying how phylogenetic and functional diversity change in function of human pressures allows us to better understand the contribution of extant lineages to ecosystem functioning in tropical forests.     

Host-ectoparasite specificity in a small mammal community in an area of Atlantic Rain Forest (Ilha Grande, State of Rio de Janeiro), Southeastern Brazil

The analyses of the ectoparasite species associated with a small mammal community on Ilha Grande, a coastal island in southern of the state of Rio de Janeiro, Brazil, evaluated the level of host-ectoparasite specificity. Was used the Jaccard index for qualitative data to analyse the similarity. The lowest value of similarity occurred between Proechimys iheringi and Marmosops incanus and between Sciurus aestuans and Nectomys squamipes (Cj=0.08) and the highest between P. iheringi and Oxymycterus sp. (Cj=0.33). This index showed a low value of similarity across the ectoparasite community. The only exception from this pattern of high host specificity occurred with P. iheringi and Oxymycterus sp., which shared five species of ectoparasites. The similarity values, for most of the cases, is smaller than 0.2.     

Characterization of SPACR, a sialoprotein associated with cones and rods present in the interphotoreceptor matrix of the human retina: immunological and lectin binding analysis

Rod and cone photoreceptors project from the outer retinal surface into a carbohydrate-rich interphotoreceptor matrix (IPM). Unique IPM glycoconjugates are distributed around rods and cones. Wheat germ agglutinin (WGA) strongly decorates the rod matrix domains and weakly decorates the cone matrix domains. This study characterizes the major WGA-binding glycoprotein in the human IPM, which we refer to as SPACR (sialoprotein associated with cones and rods). SPACR, which has a molecular weight of 147 kDa, was isolated and purified from the IPM by lectin affinity chromatography. A polyclonal antibody to SPACR was prepared that colocalizes in tissue preparations with WGA-binding domains in the IPM. Sequential digestion of SPACR with N- and O- glycosidases results in a systematic increase in electrophorectic mobility, indicating the presence of both N- and O-linked glycoconjugates. Complete deglycosylation results in a reduction in the relative molecular mass of SPACR by about 30%. Analysis of lectin binding allowed us to identify some of the structural characteristics of SPACR glycoconjugates. Treatment with neuraminidase exposes Galbeta1- 3GalNAc disaccharide as indicated by positive peanut agglutinin (PNA) staining, accompanied by the loss of WGA staining. Maackia amurensis agglutinins (MAA-1 and MAA-2), specific for sialic acid in alpha2-3 linkage to Gal, bind SPACR, while Sambucus nigra agglutinin (SNA), specific for alpha2-6 linked sialic acid, does not, indicating that the dominant glycoconjugate determinant on SPACR is the O-linked carbohydrate, NeuAcalpha2-3Galbeta1-3GalNAc. The abundance of sialic acid in SPACR suggests that this glycoprotein may contribute substantially to the polyanionic nature of the IPM. The carbohydrate chains present on SPACR could also provide sites for extensive crosslinking and participate in the formation of the ordered IPM lattice that surrounds the elongate photoreceptors projecting from the outer retinal surface.