Determination of sucrose in equine serum using liquid chromatography-mass spectrometry (LC/MS)

Mucosal integrity may be objectively assessed by determination of the absorption of exogenous substances such as sucrose. Gas chromatography-mass spectrometry (GC/MS) and liquid chromatography-mass spectrometry (LC/MS) have been reported for the accurate quantification of low concentrations of sucrose in serum. LC/MS offered the advantage of high sensitivity and mass selectivity without the need for extensive sample derivatization required for GC/MS methods. However, the high polarity and non-volatile nature of the sucrose molecule renders LC/MS techniques challenging. Previously published reports lacked sufficient detail to permit replication of methodology. Problems encountered with existing protocols included poor peak resolution and weak fragmentation of the parent molecule. This communication describes a LC/MS protocol developed to provide improved resolution and product detection.     

Structural Basis of Mycobacterial Inhibition by Cyclomarin A

Cyclomarin A (CymA) was identified as a mycobactericidal compound targeting ClpC1. However, the target was identified based on pulldown experiments and in vitro binding data, without direct functional evidence in mycobacteria. Here we show that CymA specifically binds to the N-terminal domain of ClpC1. In addition we have determined the co-crystal structure of CymA bound to the N-terminal domain of ClpC1 to high resolution. Based on the structure of the complex several mutations were engineered into ClpC1, which showed reduced CymA binding in vitro. The ClpC1 mutants were overexpressed in mycobacteria and two showed resistance to CymA, providing the first direct evidence that ClpC1 is the target of CymA. Phe⁸⁰ is important in vitro and in cells for the ClpC1-CymA interaction and this explains why other bacteria are resistant to CymA. A model for how CymA binding to the N-terminal domain of ClpC1 leads to uncontrolled proteolysis by the associated ClpP protease machinery is discussed.     

The first confirmed cases of full albinism in rajid species

Three albino skate specimens (Rajidae) were captured from the North Sea and English Channel between 2008 and 2011. Using DNA barcoding (COI gene) and morphometric analyses, species were identified as a spotted ray Raja montagui, a blonde ray Raja brachyura and a thornback ray Raja clavata. This finding represents the first record of full albinism (a lack of skin and retinal pigmentation) in rajid species.     

Prolonged P300 latency in children with the D2 dopamine receptor A1 allele.

Previous studies have indicated the presence of a hereditary component in the generation of the P300, or P3, a late positive component of the event-related potential. Moreover, the dopaminergic system has been implicated in the P3. In the present study, 98 healthy Caucasian boys, mean age of 12.5 years and of above-average intelligence, were studied. The sample was composed of 32 sons of active alcoholic (SAA) fathers, 36 sons of recovering alcoholic (SRA) fathers, and 30 sons of social drinker (SSD) fathers, with none of them having yet begun to consume alcohol or other drugs. TaqI A D2 dopamine receptor alleles (A1 and A2) were determined. A significant difference in the frequency of the A1 allele was found among these three groups of boys, with the SAA group having the highest A1 allele frequency (.313), followed by the SRA (.139) and the SSD (.133) groups. The relationship of the A1 and A2 alleles to P3 amplitude and latency was also determined. The results showed no significant difference in P3 amplitude between boys with the A1 and A2 allele. However, P3 latency was significantly longer in the total sample of boys with the A1 allele compared with those carrying the A2 allele. These findings suggest that polymorphism of the D2 dopamine receptor gene is an important determinant of P3 latency.     

Effect of relative humidity on fungal colonization of fiberglass insulation.

Fiberglass duct liners and fiberglass duct boards from eight buildings whose occupants complained of unacceptable or moldy odors in the air were found to be heavily colonized by fungi, particularly by Aspergillus versicolor. Unused fiberglass was found to be susceptible to fungal colonization in environmental chambers dependent upon relative humidity. No colonization was observed at relative humidities below 50%.     

Proton Nuclear Magnetic Resonance Spectroscopy of Primary Cells Derived from Nervous Tissue

Abstract: Cell culture techniques, high-resolution in vitro ¹H nuclear magnetic resonance (NMR) spectroscopy, and chromatographic analyses were used to compare the properties of purified cell populations derived from the PNS and cortical neurones. Cell cultures were immunocytochemically characterised with specific antibodies to ensure purity of the individual cultures. Spectra of perchloric acid extracts of cultured Schwann cells, perineural fibroblasts, dorsal root ganglion neurones, and cortical neurones displayed several common features. However, statistically significant differences were found by ¹H NMR spectroscopy in most metabolites among the cell types studied. In addition, cells could be distinguished by the presence or absence of certain amino acids. For example, N -acetylaspartate was present in dorsal root ganglion neurones and cortical neurones, γ-aminobutyric acid was present in large amounts in cortical neurones, and Schwann cell spectra displayed a large signal from glycine. These results extend our earlier findings that different cell types of the CNS exhibit highly characteristic metabolite profiles to now include the major cell types of the PNS. These latter cell types also exhibit characteristic metabolite compositions, such that even Schwann cells and oligodendrocyte type 2 astrocyte (O-2A) progenitor cells—precursors of the myelinating cells of the CNS and PNS, respectively—can be readily distinguished from each other.     

Virus Decay and Its Causes in Coastal Waters

Recent evidence suggests that viruses play an influential role within the marine microbial food web. To understand this role, it is important to determine rates and mechanisms of virus removal and degradation. We used plaque assays to examine the decay of infectivity in lab-grown viruses seeded into natural seawater. The rates of loss of infectivity of native viruses from Santa Monica Bay and of nonnative viruses from the North Sea in the coastal seawater of Santa Monica Bay were determined. Viruses were seeded into fresh seawater that had been pretreated in various ways: filtration with a 0.2-(mu)m-pore-size filter to remove organisms, heat to denature enzymes, and dissolved organic matter enrichment to reconstitute enzyme activity. Seawater samples were then incubated in full sunlight, in the dark, or under glass to allow partitioning of causative agents of virus decay. Solar radiation always resulted in increased rates of loss of virus infectivity. Virus isolates which are native to Santa Monica Bay consistently degraded more slowly in full sunlight in untreated seawater (decay ranged from 4.1 to 7.2% h(sup-1)) than nonnative marine bacteriophages which were isolated from the North Sea (decay ranged from 6.6 to 11.1% h(sup-1)). All phages demonstrated susceptibility to degradation by heat-labile substances, as heat treatment reduced the decay rates to about 0.5 to 2.0% h(sup-1) in the dark. Filtration reduced decay rates by various amounts, averaging 20%. Heat-labile, high-molecular-weight dissolved material (>30 kDa, probably enzymes) appeared responsible for about 1/5 of the maximal decay. Solar radiation was responsible for about 1/3 to 2/3 of the maximal decay of nonnative viruses and about 1/4 to 1/3 of that of the native viruses, suggesting evolutionary adaptation to local light levels. Our results suggest that sunlight is an important contributing factor to virus decay but also point to the significance of particles and dissolved substances in seawater.     

Choosing non‐redundant representative subsets of protein sequence data sets using submodular optimization

Selecting a non‐redundant representative subset of sequences is a common step in many bioinformatics workflows, such as the creation of non‐redundant training sets for sequence and structural models or selection of “operational taxonomic units” from metagenomics data. Previous methods for this task, such as CD‐HIT, PISCES, and UCLUST, apply a heuristic threshold‐based algorithm that has no theoretical guarantees. We propose a new approach based on submodular optimization. Submodular optimization, a discrete analogue to continuous convex optimization, has been used with great success for other representative set selection problems. We demonstrate that the submodular optimization approach results in representative protein sequence subsets with greater structural diversity than sets chosen by existing methods, using as a gold standard the SCOPe library of protein domain structures. In this setting, submodular optimization consistently yields protein sequence subsets that include more SCOPe domain families than sets of the same size selected by competing approaches. We also show how the optimization framework allows us to design a mixture objective function that performs well for both large and small representative sets. The framework we describe is the best possible in polynomial time (under some assumptions), and it is flexible and intuitive because it applies a suite of generic methods to optimize one of a variety of objective functions.