Induction of Prophages by Fluoroquinolones in Streptococcus pneumoniae: Implications for Emergence of Resistance in Genetically-Related Clones

Antibiotic resistance in Streptococcus pneumoniae has increased worldwide by the spread of a few clones. Fluoroquinolone resistance occurs mainly by alteration of their intracellular targets, the type II DNA topoisomerases, which is acquired either by point mutation or by recombination. Increase in fluoroquinolone-resistance may depend on the balance between antibiotic consumption and the cost that resistance imposes to bacterial fitness. In addition, pneumococcal prophages could play an important role. Prophage induction by fluoroquinolones was confirmed in 4 clinical isolates by using Southern blot hybridization. Clinical isolates (105 fluoroquinolone-resistant and 160 fluoroquinolone-susceptible) were tested for lysogeny by using a PCR assay and functional prophage carriage was studied by mitomycin C induction. Fluoroquinolone-resistant strains harbored fewer inducible prophages (17/43) than fluoroquinolone-susceptible strains (49/70) (P = 0.0018). In addition, isolates of clones associated with fluoroquinolone resistance [CC156 (3/25); CC63 (2/20), and CC81 (1/19)], had lower frequency of functional prophages than isolates of clones with low incidence of fluoroquinolone resistance [CC30 (4/21), CC230 (5/20), CC62 (9/21), and CC180 (21/30)]. Likewise, persistent strains from patients with chronic respiratory diseases subjected to fluoroquinolone treatment had a low frequency of inducible prophages (1/11). Development of ciprofloxacin resistance was tested with two isogenic strains, one lysogenic and the other non-lysogenic: emergence of resistance was only observed in the non-lysogenic strain. These results are compatible with the lysis of lysogenic isolates receiving fluoroquinolones before the development of resistance and explain the inverse relation between presence of inducible prophages and fluoroquinolone-resistance.     

Phylogenetic relationships and evolution of pulmonate gastropods (Mollusca): new insights from increased taxon sampling

Phylogenetic relationships among higher clades of pulmonate gastropods are reconstructed based on a data set including one nuclear marker (complete ribosomal 18S) and two mitochondrial markers (partial ribosomal 16S and Cytochrome oxidase I) for a total of 96 species. Sequences for 66 of these species are new to science, with a special emphasis on sampling the Ellobiidae, Onchidiidae, and Veronicellidae. Important results include the monophyly of Systellommatophora (Onchidiidae and Veronicellidae) as well as the monophyly of Ellobiidae (including Trimusculus, Otina, and Smeagol). Relationships within Ellobiidae, Onchidiidae, and Veronicellidae are evaluated here for the first time using molecular data. Present results are compared with those from the recent literature, and the current knowledge of phylogenetic relationships among pulmonate gastropods is reviewed: despite many efforts, deep nodes are still uncertain. Identification uncertainties about early fossils of pulmonates are reviewed. Impacts of those phylogenetic and fossil record uncertainties on our understanding of the macro-evolutionary history of pulmonates, especially transitions between aquatic and terrestrial habitats, are discussed.     

PSysCal: a parallel tool for calibration of ecosystem models

The methods used for ecosystem modelling are generally based on differential equations. Nowadays, new computational models based on concurrent processing of multiple agents (multi-agents) or the simulation of biological processes with the Population Dynamic P-System models (PDPs) are gaining importance. These models have significant advantages over traditional models, such as high computational efficiency, modularity and its ability to model the interaction between different biological processes which operate concurrently. By this, they are becoming useful for simulating complex dynamic ecosystems, untreatable with classical techniques. On the other hand, the main counterpart of P-System models is the need for calibration. The model parameters represent the field measurements taken by experts. However, the exact values of some of these parameters are unknown and experts define a numerical interval of possible values. Therefore, it is necessary to perform a calibration process to fit the best value of each interval. When the number of unknown parameters increases, the calibration process becomes computationally complex and storage requirements increase significantly. In this paper, we present a parallel tool (PSysCal) for calibrating next generation PDP models. The results shown that the calibration time is reduced exponentially with the amount of computational resources. However, the complexity of the calibration process and a limitation in the number of available computational resources make the calibration process intractable for large models. To solve this, we propose a heuristic technique (PSysCal+H). The results show that this technique significantly reduces the computational cost, it being practical for solving large model instances even with limited computational resources.     

Effect of CRF injected into the median eminence on GH secretion in female rats under different steroid status

To evaluate whether the median eminence (ME) is a site of action of CRF (corticotropin releasing factor) on GH secretion and to determine the possible role of estradiol and progesterone in modifying theses secretion, we injected CRF (0.25, 0.75, 1, and 1.5 nmol of peptide dissolved in 1 μl of water) directly into the ME in three experimental groups of rats: Long-term ovariectomized (OVX); OVX primed by estradiol (OVX±E) and OVX primed by estradiol plus progesterone (OVX±EP). Blood was collected to determine GH (30, 60, 90, and 120 min postinjection) Serum T₃, T₄, and glucose levels were measured in OVX±E rats 30 min postinjection. CRF at all doses studied significantly decreased serum GH levels in the three experimental groups. Serum T₃, T₄, and glucose levels were unchanged after CRF administration. The results suggest that: CRF inhibits “per se” GH secretion, at least in part, by a central action in the ME. The inhibitory effect of CRF on GH is independent of the estrogen/progesterone status of the animal. CRF at ME levels may participate in a variety of stress-related responses, including growth inhibition, through GH suppression.     

Crystal structures of Xanthomonas campestris OleA reveal features that promote head-to-head condensation of two long-chain fatty acids

OleA is a thiolase superfamily enzyme that has been shown to catalyze the condensation of two long-chain fatty acyl-coenzyme A (CoA) substrates. The enzyme is part of a larger gene cluster responsible for generating long-chain olefin products, a potential biofuel precursor. In thiolase superfamily enzymes, catalysis is achieved via a ping-pong mechanism. The first substrate forms a covalent intermediate with an active site cysteine that is followed by reaction with the second substrate. For OleA, this conjugation proceeds by a nondecarboxylative Claisen condensation. The OleA from Xanthomonas campestris has been crystallized and its structure determined, along with inhibitor-bound and xenon-derivatized structures, to improve our understanding of substrate positioning in the context of enzyme turnover. OleA is the first characterized thiolase superfamily member that has two long-chain alkyl substrates that need to be bound simultaneously and therefore uniquely requires an additional alkyl binding channel. The location of the fatty acid biosynthesis inhibitor, cerulenin, that possesses an alkyl chain length in the range of known OleA substrates, in conjunction with a single xenon binding site, leads to the putative assignment of this novel alkyl binding channel. Structural overlays between the OleA homologues, 3-hydroxy-3-methylglutaryl-CoA (HMG-CoA) synthase and the fatty acid biosynthesis enzyme FabH, allow assignment of the two remaining channels: one for the thioester-containing pantetheinate arm and the second for the alkyl group of one substrate. A short β-hairpin region is ordered in only one of the crystal forms, and that may suggest open and closed states relevant for substrate binding. Cys143 is the conserved catalytic cysteine within the superfamily, and the site of alkylation by cerulenin. The alkylated structure suggests that a glutamic acid residue (Glu117β) likely promotes Claisen condensation by acting as the catalytic base. Unexpectedly, Glu117β comes from the other monomer of the physiological dimer.     

A diverse Upper Jurassic dinosaur ichnofauna from central‐west Portugal

Mateus, O. & Milàn, J. 2009: A diverse Upper Jurassic dinosaur ichnofauna from central‐west Portugal. Lethaia, Vol. 43, pp. 245–257. A newly discovered dinosaur track‐assemblage from the Upper Jurassic Lourinhã Formation (Lusitanian Basin, central‐west Portugal), comprises medium‐ to large‐sized sauropod tracks with well‐preserved impressions of soft tissue anatomy, stegosaur tracks and tracks from medium‐ to large‐sized theropods. The 400‐m‐thick Lourinhã Formation consists of mostly aluvial sediments, deposited during the early rifting of the Atlantic Ocean in the Kimmeridgian and Tithonian. The stratigraphic succession shows several shifts between flood‐plain mud and fluvial sands that favour preservation and fossilization of tracks. The studied track‐assemblage is found preserved as natural casts on the underside of a thin bivalve‐rich carbonate bed near the Tithonian–Kimmeridgian boundary. The diversity of the tracks from the new track assemblage is compared with similar faunas from the Upper Jurassic of Asturias, Spain and the Middle Jurassic Yorkshire Coast of England. The Portuguese record of Upper Jurassic dinosaur body fossils show close similarity to the track fauna from the Lourinhã Formation. □Dinosaur tracks, Lusitanian Basin, Portugal, skin impressions, Upper Jurassic.     

Toward an early marker of metabolic dysfunction: omentin-1 in prepubertal children

Omentin-1 is a recently recognized adipokine primarily originating in visceral adipose tissue. We posited that circulating omentin-1 could be an early marker of metabolic dysfunction. To this end, we examined the associations between circulating omentin-1, body fat (bioelectric impedance), an endocrine-metabolic profile (homeostasis model assessment for insulin resistance (HOMA(IR)), serum lipids, high-molecular-weight (HMW) adiponectin and blood pressure (BP)) and family history of obesity and diabetes in asymptomatic prepubertal children (n = 161; 77 boys and 84 girls; age 7 ± 1 year) with a normal distribution of height and weight. Increased circulating omentin-1 was associated with a poorer metabolic profile, with relatively higher HOMA(IR), fasting triacylglycerol, BP and familial prevalence of diabetes (all P < 0.005 to P < 0.0001), and relatively lower fraction of HMW adiponectin (P < 0.005), whereas no relationship was found with body weight or fat or with family history of obesity. All these associations were independent of age, gender and fat mass. In conclusion, circulating omentin-1 may become a marker of metabolic dysfunction integrating insulin sensitivity, markers of adipose-tissue metabolism and BP as early as in prepubertal childhood.     

Lung squamous cell carcinoma and adenocarcinoma cell lines use different mediators to induce comparable phenotypic and functional changes in human monocyte-derived dendritic cells

Tumor-derived immunosuppressive factors contribute to the evasion of malignant cells from the immune response, partially by hampering dendritic cell (DC) differentiation. Here, we analyze whether soluble mediators released by the most frequent histological types of non-small cell lung carcinoma, squamous cell carcinoma (SCC), and adenocarcinoma (AD) cells, affect the development and functionality of DC. Monocytes from healthy donors were differentiated in vitro into DC with granulocyte–macrophage colony-stimulating factor (GM-CSF) and interleukin (IL)-4, in the absence or presence of soluble factors (SF) from SCC or AD cell lines. Monocytes were differentiated in parallel into macrophages (MΦ s) with macrophage colony-stimulating factor (M-CSF). SF-treated DC were phenotypically and functionally more similar to MΦ s than to untreated DC [control DC (Ctrl-DC)]. Both tumors increased myelomonocytic markers (CD14, CD16, CD32, and CD163) and impaired CD1a expression on DC. SF-treated DC increased their endocytic capacity, and released higher levels of IL-6, IL-10, and lower levels of IL-12, compared to Ctrl-DC. SF-treated DC were poor stimulators in mixed lymphocyte reactions, and naïve CD4⁺ T lymphocytes stimulated by SF-treated DC secreted lower levels of interferon (IFN)-γ and higher amounts of IL-10 than controls. In contrast to AD, the effects caused by SCC were mostly abolished by IL-6 neutralization during monocyte differentiation. However, tumor-derived prostanoid blockade recovered the IFN-γ levels secreted by lymphocytes stimulated with SF-treated DC, whereas prostanoid/IL-6 or prostanoid/IL-10 blockade decreased IL-10 production only by SCC-DC-stimulated lymphocytes. Thus, we provide evidence that lung SCC and AD cause comparable deficiencies on DC in vitro, skewing monocyte differentiation from DC to MΦ -like cells, but most of these changes occurred via different mediators.