An amphipathic cyclic tetrapeptide scaffold containing halogenated β2,2‐amino acids with activity against multiresistant bacteria

The present study describes the synthesis and biological studies of a small series of head‐to‐tail cyclic tetrapeptides of the general structure c(Lys‐β^2,2‐Xaa‐Lys) containing one lipophilic β^2,2‐amino acid and Lys, Gly, Ala, or Phe as the Xaa residue in the sequence. The peptides were investigated for antimicrobial activity against gram‐positive and gram‐negative reference strains and 30 multiresistant clinical isolates including strains with extended spectrum β‐lactamase—carbapenemase (ESBL‐CARBA) production. Toxicity was determined against human red blood cells. The most potent peptides showed high activity against the gram‐positive clinical isolates with minimum inhibitory concentrations of 4–8 μg/mL and low haemolytic activity. The combination of high antimicrobial activity and low toxicity shows that these cyclic tetrapeptides containing lipophilic β^2,2‐amino acids form a valuable scaffold for designing novel antimicrobial agents.     

Life cycle of hake and likely management implications

Despite its economic and social importance for Namibia and South Africa, limited documented information exists regarding key aspects of the biology of deep-water hake, including its life cycle. This study utilizes data collected through the demersal surveys of the R/V Dr Fridtjof Nansen in South Africa and F/V Blue Sea 1 in Namibia to describe the migratory patterns of deep-water hake in space and time. Furthermore the study investigates aspects of the life cycle of this important species in the Benguela region. Results show that deep-water hake spawns between the western Agulhas Bank and Elands Bay in South Africa with the main nursery ground between Hondeklip Bay and the northern tip of Orange Banks. Deep-water hake in Namibia (up to the Kunene River) and along the south coast of South Africa (eastwards to Port Alfred) originate from these grounds, and undertake long-range migrations across latitudes and longitudes, respectively. This hypothesis is supported by the finding that spawning has not been observed in Namibia and there are no small juveniles along the South African south coast from the eastern border of the Agulhas Bank. The proposed pattern implies an interconnection between the Namibian and the South African components of the stock and the consequent need for a revision of the present management regime based on the assumption of stocks confined within the respective national jurisdictions. This study has used length frequency distributions in space and time in order to investigate the life cycle, in terms of origin, movement and population structure in particular, an approach that may also be useful for other widely distributed species.     

Localization of monoaminergic neurons in the central nervous system of Astacus astacus Linné (Crustacea)

Summary The cellular localization of biogenic monoamines in crustaceans was studied by means of a highly specific and sensitive fluorescence method devised by Falck and Hillarp. It was found that neurons displaying specific fluorescence in the central nervous system were confined to the protocerebrum, the medulla externa and interna and the ventral nerve cord. The method allows a distinction between the fluorophores of 5-hydroxytryptamine (and 5-hydroxytryptophan), which emit the yellow light, and the fluorophores deriving from the catecholamines (and DOPA), which emit the green light. Green-fluorescent neurons occurred abundantly in the aforementioned parts of the central nervous system while yellow-fluorescent neurons were sparsely present in the same parts.The present work has been carried out at the departments of Histology and Zoology at the University of Lund. The authors take great pleasure in expressing their warmest thanks for laboratory facilities, provided by Professors Erik Dahl (Zoological Institute) and Bengt Falck (Histological Institute).The research reported in this document has been sponsored by the Air Force Office of Scientific Research under Grant AF EOAR 66-14 through the European Office of Aerospace Research (OAR), United States Air Force and by a grant from the Swedish Natural Science Research Council 99-32 (nr 5995).     

Infection with human cytomegalovirus alters the MMP-9/TIMP-1 balance in human macrophages

Human cytomegalovirus (HCMV) has been suggested to contribute to the development of vascular diseases. Since matrix metalloproteinases (MMPs) have been implicated in atherosclerosis and plaque rupture, we investigated the effect of HCMV infection on MMP expression in human macrophages. We used quantitative real-time PCR, Western blotting, and gelatin zymography to study the expression and activity of MMP-2, -3, -7, -9, -12, -13, and -14 and of tissue inhibitor of metalloproteinase 1 (TIMP-1), -2, -3, and -4. HCMV infection reduced MMP-9 mRNA, protein, and activity levels but increased TIMP-1 mRNA and protein levels. Furthermore, a decrease in MMP-12, MMP-14, TIMP-2, and TIMP-3 mRNA levels could be detected. The MMP-9 and TIMP-1 mRNA alterations required viral replication. MMP-9 mRNA expression was affected by an immediate-early or early viral gene product, whereas TIMP-1 mRNA expression was affected by late viral gene products. We conclude that HCMV infection specifically alters the MMP-9/TIMP-1 balance in human macrophages, which in turn reduces MMP-9 activity in infected cells. Since MMP-9 prevents atherosclerotic plaque development in mice, these results suggest that HCMV may contribute to atherogenesis through specific effects on MMP-9 activity.     

Identification of a core set of genes that signifies pathways underlying cardiac hypertrophy

Although the molecular signals underlying cardiac hypertrophy have been the subject of intense investigation, the extent of common and distinct gene regulation between different forms of cardiac hypertrophy remains unclear. We hypothesized that a general and comparative analysis of hypertrophic gene expression, using microarray technology in multiple models of cardiac hypertrophy, including aortic banding, myocardial infarction, an arteriovenous shunt and pharmacologically induced hypertrophy, would uncover networks of conserved hypertrophy-specific genes and identify novel genes involved in hypertrophic signalling. From gene expression analyses (8740 probe sets, n = 46) of rat ventricular RNA, we identified a core set of 139 genes with consistent differential expression in all hypertrophy models as compared to their controls, including 78 genes not previously associated with hypertrophy and 61 genes whose altered expression had previously been reported. We identified a single common gene program underlying hypertrophic remodelling, regardless of how the hypertrophy was induced. These genes constitute the molecular basis for the existence of one main form of cardiac hypertrophy and may be useful for prediction of a common therapeutic approach. Supplementary material for this article can be found at: http://www.interscience.wiley.com/jpages/1531-6912/suppmat.     

Effects of Ebola virus glycoproteins on endothelial cell activation and barrier function

Ebola virus causes severe hemorrhagic fever with high mortality rates in humans and nonhuman primates. Vascular instability and dysregulation are disease-decisive symptoms during severe infection. While the transmembrane glycoprotein GP(1,2) has been shown to cause endothelial cell destruction, the role of the soluble glycoproteins in pathogenesis is largely unknown; however, they are hypothesized to be of biological relevance in terms of target cell activation and/or increase of endothelial permeability. Here we show that virus-like particles (VLPs) consisting of the Ebola virus matrix protein VP40 and GP(1,2) were able to activate endothelial cells and induce a decrease in barrier function as determined by impedance spectroscopy and hydraulic conductivity measurements. In contrast, the soluble glycoproteins sGP and delta-peptide did not activate endothelial cells or change the endothelial barrier function. The VLP-induced decrease in barrier function was further enhanced by the cytokine tumor necrosis factor alpha (TNF-alpha), which is known to induce a long-lasting decrease in endothelial cell barrier function and is hypothesized to play a key role in Ebola virus pathogenesis. Surprisingly, sGP, but not delta-peptide, induced a recovery of endothelial barrier function following treatment with TNF-alpha. Our results demonstrate that Ebola virus GP(1,2) in its particle-associated form mediates endothelial cell activation and a decrease in endothelial cell barrier function. Furthermore, sGP, the major soluble glycoprotein of Ebola virus, seems to possess an anti-inflammatory role by protecting the endothelial cell barrier function.     

Milk production and composition in reindeer (Rangifer tarandus): effect of lactational stage

Milk yield and composition of major milk constituents were measured in captive, nursing reindeer. Registration of milk production was performed during two successive lactations (2001 and 2002). The milk yield was significantly affected by week of lactation (P<0.001) and by individual (P<0.001). The lactation curve had an asymmetrical peak 3 weeks postpartum and the milk yield at peak lactation was 983 g/day (range 595-1239). The length of lactation varied from 24 to 26 weeks and average total milk production was 99.5 kg. From peak lactation the milk production decreased linearly (P<0.001) until milk production was terminated. Mean values for content of major milk constituents were 15.5% fat, 9.9% protein and 2.5% lactose. The content of fat and protein increased markedly with the lactation stage (P<0.001), while lactose showed a slight decrease (P<0.001). The milk composition was significantly affected by stage of lactation (P<0.001). There was a marginally significant decrease in protein:fat ratio (P=0.06) as protein was substituted by fat with stage of lactation. The caloric value of the milk averaged 8.7 kJ/g and increased significantly with the stage of lactation (P<0.001). The overall increase in milk gross energy content during lactation was 67.6%. The energy output averaged 7996 kJ/day at peak lactation and decreased significantly during the course of lactation (P=0.002).     

Evaluation of exudates by solid phase microextraction-gas chromatography

Head-space solid phase microextraction combined with gas chromatography (SPME-GC) was used for the determination of bacterial volatile fatty acid (VFA) patterns. The method was validated with cultures of reference bacterial strains. It was confirmed that VFA production depends on the composition of the cultivation medium, which limits accurate characterisation of particular bacterial species. A set of 195 clinical exudates of various origin and consistence was analysed using SPME-GC and compared with 73 samples extracted using tert-butyl methyl ether. Approximate agreement of VFA profiles with cultivation findings was found in most cases. However, 20.5% of clinical exudates with distinct VFA profiles appeared to be false-negative by cultivation. Using SPME-GC of exudates, the frequency of false-negative cultivations was higher than that of solvent extraction of exudates or blood cultures found previously. The described method is suitable for preliminary detection of bacteria, particularly non-sporulating anaerobes, in clinical samples. It can reveal false-negative findings due to cultivation. Analysis can be performed in 30 min without the need for cultivation.     

Comparative analysis of the mitochondrial cytochrome c oxidase subunit I (COI) gene in ciliates (Alveolata,Ciliophora) and evaluation of its suitability as a biodiversity marker

The mitochondrial cytochrome c oxidase subunit 1 (COI) gene of ciliates was first successfully sequenced in species of the genera Tetrahymena and Paramecium (Class Oligohymenophorea). The sequence of the COI gene is extremely divergent from other eukaryotes and includes an insert, which is over 300 nucleotides long. In this study, we designed a primer pair that successfully amplified the COI gene of ciliates from five different classes: Heterotrichea, Spirotrichea, Oligohymenophorea, Nassophorea and Colpodea. These classes represent the diversity of the phylum Ciliophora very well, since they are widely distributed on the ciliate small subunit rRNA tree. The amplified region is approximately 850 nucleotides long and corresponds to the general barcoding region; it also includes the insert region. In this study, 58 new COI sequences from over 38 species in 13 orders are analysed and compared, and distance trees are constructed. While the COI gene shows high divergence within ciliates, the insert region, which is present in all classes, is even more divergent. Genetic distances calculated with and without the insert region remain in the same range at the intraspecific level, but they differ considerably at or above genus level. This suggests that the entire barcoding region is under similar selective constraints and that the evolutionary rate of the ciliate COI is extremely high and shows unequal rate variation. Although many problems still remain regarding standardization of barcoding methods in ciliates, the development of a universal or almost universal primer combination for the Phylum Ciliophora represents important progress. As shown in four examples, the resolution of COI at the intraspecific level is much greater than that of any nuclear genes and shows great potential to (1) identify species based on molecular data if a reliable database exists, and (2) resolve the relationships of closely related ciliate taxa and uncover cryptic species.