Molecular diet analysis of two african free-tailed bats (molossidae) using high throughput sequencing

Given the diversity of prey consumed by insectivorous bats, it is difficult to discern the composition of their diet using morphological or conventional PCR-based analyses of their faeces. We demonstrate the use of a powerful alternate tool, the use of the Roche FLX sequencing platform to deep-sequence uniquely 5' tagged insect-generic barcode cytochrome c oxidase I (COI) fragments, that were PCR amplified from faecal pellets of two free-tailed bat species Chaerephon pumilus and Mops condylurus (family: Molossidae). Although the analyses were challenged by the paucity of southern African insect COI sequences in the GenBank and BOLD databases, similarity to existing collections allowed the preliminary identification of 25 prey families from six orders of insects within the diet of C. pumilus, and 24 families from seven orders within the diet of M. condylurus. Insects identified to families within the orders Lepidoptera and Diptera were widely present among the faecal samples analysed. The two families that were observed most frequently were Noctuidae and Nymphalidae (Lepidoptera). Species-level analysis of the data was accomplished using novel bioinformatics techniques for the identification of molecular operational taxonomic units (MOTU). Based on these analyses, our data provide little evidence of resource partitioning between sympatric M. condylurus and C. pumilus in the Simunye region of Swaziland at the time of year when the samples were collected, although as more complete databases against which to compare the sequences are generated this may have to be re-evaluated.     

Three-dimensional structure of the rSly1 N-terminal domain reveals a conformational change induced by binding to syntaxin 5

Sec1/Mun18-like (SM) proteins and soluble N-ethylmaleimide-sensitive factor attachment protein receptors (SNAREs) play central roles in intracellular membrane fusion. Diverse modes of interaction between SM proteins and SNAREs from the syntaxin family have been described. However, the observation that the N-terminal domains of Sly1 and Vps45, the SM proteins involved in traffic at the endoplasmic reticulum, the Golgi, the trans-Golgi network and the endosomes, bind to similar N-terminal sequences of their cognate syntaxins suggested a unifying theme for SM protein/SNARE interactions in most internal membrane compartments. To further understand this mechanism of SM protein/SNARE coupling, we have elucidated the structure in solution of the isolated N-terminal domain of rat Sly1 (rSly1N) and analyzed its complex with an N-terminal peptide of rat syntaxin 5 by NMR spectroscopy. Comparison with the crystal structure of a complex between Sly1p and Sed5p, their yeast homologues, shows that syntaxin 5 binding requires a striking conformational change involving a two-residue shift in the register of the C-terminal beta-strand of rSly1N. This conformational change is likely to induce a significant alteration in the overall shape of full-length rSly1 and may be critical for its function. Sequence analyses indicate that this conformational change is conserved in the Sly1 family but not in other SM proteins, and that the four families represented by the four SM proteins found in yeast (Sec1p, Sly1p, Vps45p and Vps33p) diverged early in evolution. These results suggest that there are marked distinctions between the mechanisms of action of each of the four families of SM proteins, which may have arisen from different regulatory requirements of traffic in their corresponding membrane compartments.     

Axenic promastigote forms of Leishmania (Viannia) lainsoni as an alternative source for Leishmania antigen production

The present study demonstrates that axenic cultures of Leishmania (Viannia) lainsoni produce larger cell masses in NNN-LIT medium, as well as higher amounts of total proteins in cell extracts, than Leishmania (Leishmania) amazonensis. Antigenicity of L. (V.) lainsoni whole promastigotes is similar to that of L. (L.) amazonensis, as demonstrated by an indirect immunofluorescence diagnostic test using sera from human patients and dogs infected with visceral leishmaniasis. Infectivity of the L. (V.) lainsoni strain used in the present work was demonstrated by the detection by transmission-electron microscopy of tissue amastigotes in skin lesion samples from an experimentally infected hamster. Incubation of lesion fragments in NNN-LIT medium allowed us to obtain promastigote forms, which could be cultivated successfully in vitro. lsoenzyme analysis of such promastigotes confirmed the parasite strain as L. (V.) lainsoni, as compared to other Leishmania reference strains. Our data indicate that L. (V.) lainsoni is a useful alternative source for antigen production as well for use in assays that depend on large cell volumes of Leishmania spp. parasites.     

The finding of Echinostoma (Trematoda: Digenea) and hookworm eggs in coprolites collected from a Brazilian mummified body dated 600-1,200 years before present

The identification of parasites from ancient cultures expands our list of parasites infective to extant humans. A partially mummified human body from the archeological site of Lapa do Boquete, Minas Gerais State, Brazil, was recently discovered. It was interred between 600 and 1,200 yr ago. Dietary analysis showed that the mummified body was from a society that had a mixed subsistence of agriculture and gathering of wild foods. Coprolites from the body contained numerous helminth eggs. The eggs were identified as those of Echinostoma sp. and hookworm. Hookworm infection in pre-Columbian populations is already established, but this is the first evidence of Echinostoma sp. eggs found in human coprolites. The diagnosis of a true infection, as opposed to false parasitism, is discussed. The possibility of Echinostoma ilocanum infection is discussed, as this is a common species found in humans in the Asiatic region, which could have been introduced in South America in the pre-Columbian period. Alternative possibilities are also considered, including indigenous Brazilian Echinostoma species.     

Role of serotonin receptors in panic-like behavior induced by nitric oxide in the rat dorsolateral periaqueductal gray: effects of chronic clomipramine treatment

Local administration of serotonin (5-HT) receptor agonists inhibits panic-like reactions induced by electrical stimulation of the rat dorsolateral periaqueductal grey (dlPAG). This anti-aversive effect is enhanced by chronic treatment with anti-panic drugs such as clomipramine. Since nitric oxide (NO) may mediate panic-like behavior in the dlPAG, we tested the hypothesis that chronic clomipramine treatment would also potentiate the effects of locally injected 5-HT-receptor agonists on panic-like reactions induced by intra-dlPAG injection of an NO-donor (SIN). After 21 days of daily i.p. injections of saline or clomipramine (10 mg/kg) male Wistar rats received local injections of saline, the 5-HT(1A)-receptor agonist 8-OH-DPAT (8 nmol) or the 5-HT2A/2C-receptor agonist DOI (16 nmol) followed by saline or SIN (150 nmol). NO-induced panic-like reactions were inhibited by DOI, but not by 8-OH-DPAT. Chronic clomipramine did not modify these effects but tended to produce anti-aversive effect by itself. In chronically clomipramine treated animals 8-OH-DPAT potentiated NO-induced panic-like reactions. The results indicate that the panic-like effects of NO in the dlPAG may be attenuated by 5-HT2A/2C-, but not by 5-HT1A-receptors. The anti-aversive effect of DOI is not modified by chronic clomipramine treatment.     

Synthesis of benzoyl phenyl benzoates as effective inhibitors for phospholipase A2 and hyaluronidase enzymes

Benzoylation of (hydroxy phenyl) phenyl methanone 2a-g to benzoyl phenyl benzoates 4a-g, a benzophenone analogue, was achieved in good yield. All the newly synthesized compounds were evaluated for their phospholipase A2 [E.C. 3.1.1.4] and hyaluronidase [E.C. 3.2.1.35] enzyme inhibitory activity in snake venom as source and their structure-activity relationship with respect to different groups is reported for the first time. The in vitro PLA2 enzyme inhibitory activity and in vivo anti-inflammatory activity studies of benzoyl phenyl benzoates are illustrated.     

Antinociceptive effects of the essential oil of Alpinia zerumbet on mice.

Alpinia zerumbet (Pers.) Burtt. et Smith is an aromatic plant that is distributed widely in the tropical and sub-tropical regions of the world. In Brazil, where A. zerumbet is called "colonia", it is used widely in folk medicine for the treatment of various diseases, including hypertension. In the present study, the antinociceptive effects of the orally administered essential oil of A. zerumbet (EOAz) were evaluated in male Swiss mice (20-25 g each). In the acetic acid-induced writhing test, EOAz (30, 100 and 300 mg/kg body wt.; n = 10, n = 13 and n = 15, respectively) was effective at all doses. In the hot-plate test, EOAz significantly increased the latency at doses of 100 and 300 mg/kg body wt., but not at 30 mg/kg body wt., at all observation times up to the 180th min (n = 10 for each dose). In the formalin test, EOAz significantly reduced paw licking time in the second phase of the test at 100 mg/kg body wt. (n = 10), but decreased it in both phases at 300 mg/kg body wt. (n = 10). At 30 mg/kg body wt., the effect of EOAz did not differ from control values in either phase of the formalin test (n = 10). Pretreatment with naloxone (5 mg/kgbodywt., i.p.) caused a significant reversal of the analgesic effect of 300 mg/kg body wt. EOAz (n = 8) that was complete for the first phase, but only partial for the second phase of the formalin test. The data show that orally administered OEAz promotes a dose-dependent antinociceptive effect, with a mechanism of action which probably involves the participation of opiate receptors.     

mummy/cystic encodes an enzyme required for chitin and glycan synthesis, involved in trachea, embryonic cuticle and CNS development--analysis of its role in Drosophila tracheal morphogenesis

Tracheal and nervous system development are two model systems for the study of organogenesis in Drosophila. In two independent screens, we identified three alleles of a gene involved in tracheal, cuticle and CNS development. Here, we show that these alleles, and the previously identified cystic and mummy, all belong to the same complementation group. These are mutants of a gene encoding the UDP-N-acetylglucosamine diphosphorylase, an enzyme responsible for the production of UDP-N-acetylglucosamine, an important intermediate in chitin and glycan biosynthesis. cyst was originally singled out as a gene required for the regulation of tracheal tube diameter. We characterized the cyst/mmy tracheal phenotype and upon histological examination concluded that mmy mutant embryos lack chitin-containing structures, such as the procuticle at the epidermis and the taenidial folds in the tracheal lumen. While most of their tracheal morphogenesis defects can be attributed to the lack of chitin, when compared to krotzkopf verkehrt (kkv) chitin-synthase mutants, mmy mutants showed a stronger phenotype, suggesting that some of the mmy phenotypes, like the axon guidance defects, are chitin-independent. We discuss the implications of these new data in the mechanism of size control in the Drosophila trachea.