Toward an optimal sampling protocol for Hemiptera on understorey plants

There are no standardised sampling protocols for inventorying Hemiptera from understorey or canopy plants. This paper proposes an optimal protocol for the understorey, after evaluating the efficiency of seven methods to maximise the richness of Hemiptera collected from plants with minimal field and laboratory time. The methods evaluated were beating, chemical knockdown, sweeping, branch clipping, hand collecting, vacuum sampling and sticky trapping. These techniques were tested at two spatial scales: 1 ha sites and individual plants. In addition, because efficiency may differ with vegetation structure, sampling of sites was conducted in three disparate understorey habitats, and sampling of individual plants was conducted across 33 plant species. No single method sampled the majority of hemipteran species in the understorey. Chemical knockdown, vacuum sampling and beating yielded speciose samples (61, 61 and 30 species, respectively, representing 53, 53 and 26% of total species collected). The four remaining methods provided species-poor samples (<18 species or <16% of total species collected). These methods also had biases towards particular taxa (e.g., branch clipping and hand collecting targeted sessile Hemiptera, and sticky trapping were dominated by five species of Psyllidae). The most time-efficient methods were beating, sweeping and hand collecting (200 minutes of field and laboratory time yielded >7 species for each technique). By comparison, vacuum sampling, sticky trapping, branch clipping and chemical knockdown yielded <5 species for the same period. Chemical knockdown had further disadvantages; high financial cost and potential spray drift. The most effective methods for a standardised sampling protocol to inventory Hemiptera from the understorey are beating and vacuum sampling. If used in combination, these methods optimise the catch of understorey hemipteran species, as their samples have high complementarity.     

Properties of the permeability transition pore in mitochondria devoid of Cyclophilin D

We have studied the properties of the permeability transition pore (PTP) in mitochondria from the liver of mice where the Ppif gene encoding for mitochondrial Cyclophilin D (CyP-D) had been inactivated. Mitochondria from Ppif-/- mice had no CyP-D and displayed a striking desensitization of the PTP to Ca2+, in that pore opening required about twice the Ca2+ load necessary to open the pore in strain-matched, wild-type mitochondria. Mitochondria lacking CyP-D were insensitive to Cyclosporin A (CsA), which increased the Ca2+ retention capacity only in mitochondria from wild-type mice. The PTP response to ubiquinone 0, depolarization, pH, adenine nucleotides, and thiol oxidants was similar in mitochondria from wild-type and Ppif-/- mice. These experiments demonstrate that (i) the PTP can form and open in the absence of CyP-D, (ii) that CyP-D represents the target for PTP inhibition by CsA, and (iii) that CyP-D modulates the sensitivity of the PTP to Ca2+ but not its regulation by the proton electrochemical gradient, adenine nucleotides, and oxidative stress. These results have major implications for our current understanding of the PTP and its modulation in vitro and in vivo.     

Disease-related prion protein forms aggresomes in neuronal cells leading to caspase activation and apoptosis

The molecular basis for neuronal death in prion disease is not established, but putative pathogenic roles for both disease-related prion protein (PrP(Sc)) and accumulated cytosolic PrP(C) have been proposed. Here we report that only prion-infected neuronal cells become apoptotic after mild inhibition of the proteasome, and this is strictly dependent upon sustained propagation of PrP(Sc). Whereas cells overexpressing PrP(C) developed cytosolic PrP(C) aggregates, this did not cause cell death. In contrast, only in prion-infected cells, mild proteasome impairment resulted in the formation of large cytosolic perinuclear aggresomes that contained PrP(Sc), heat shock chaperone 70, ubiquitin, proteasome subunits, and vimentin. Similar structures were found in the brains of prion-infected mice. PrP(Sc) aggresome formation was directly associated with activation of caspase 3 and 8, resulting in apoptosis. These data suggest that neuronal propagation of prions invokes a neurotoxic mechanism involving intracellular formation of PrP(Sc) aggresomes. This, in turn, triggers caspase-dependent apoptosis and further implicates proteasome dysfunction in the pathogenesis of prion diseases.     

Neurotransmitter release regulated by a MALS-liprin-alpha presynaptic complex

Synapses are highly specialized intercellular junctions organized by adhesive and scaffolding molecules that align presynaptic vesicular release with postsynaptic neurotransmitter receptors. The MALS/Veli-CASK-Mint-1 complex of PDZ proteins occurs on both sides of the synapse and has the potential to link transsynaptic adhesion molecules to the cytoskeleton. In this study, we purified the MALS protein complex from brain and found liprin-alpha as a major component. Liprin proteins organize the presynaptic active zone and regulate neurotransmitter release. Fittingly, mutant mice lacking all three MALS isoforms died perinatally with difficulty breathing and impaired excitatory synaptic transmission. Excitatory postsynaptic currents were dramatically reduced in autaptic cultures from MALS triple knockout mice due to a presynaptic deficit in vesicle cycling. These findings are consistent with a model whereby the MALS-CASK-liprin-alpha complex recruits components of the synaptic release machinery to adhesive proteins of the active zone.     

Prior exposure to indole-3-carbinol decreases the incidence of specific cyclophosphamide-induced developmental defects in mice

BACKGROUND: Indole-3-carbinol (I3C) is a product of the hydrolysis of glucobrassicin that is found in cruciferous vegetables. I3C can intervene in toxic processes that are mediated by oxidative mechanisms because it possesses the chemical and pharmacokinetic properties necessary to provide a free radical trap. Cyclophosphamide (CP) is a bifunctional alkylating agent known to produce DNA damage and to cause developmental toxicity, including malformations, in laboratory animals. METHODS: Pregnant CD-1 mice were given a 100 mg/kg dose of I3C 24 or 48 hr before administration of 20 mg/kg CP on gestation day 10 (GD 10). Controls were given the vehicle (DMSO), I3C, or CP. This regimen was carried out to determine if I3C could protect against the developmental toxicity of alkylating agents, such as CP. Dams were sacrificed on GD 17 and their litters were examined for adverse effects. RESULTS: Treatment with I3C 48 hr before CP administration was associated with decreased fetal limb and tail malformations. Limb malformation incidences were reduced from 42% litters affected in the CP control to 16% in the I3C/CP 48-hr treatment group, and tail malformations were reduced from 45% in the CP control to 16% in the I3C/CP 48-hr treatment group, indicating a protective effect of prior exposure to I3C. I3C given 24 hr before CP had no significant protective effect, while having an apparently adverse consequence with regard to the incidence of talipes. CONCLUSIONS: Exposure of a developing mammal to indole-3-carbinol before exposure to cyclophosphamide during organogenesis can influence the teratogenicity of cyclophosphamide.     

Endogenous Nmnat2 Is an Essential Survival Factor for Maintenance of Healthy Axons

The molecular triggers for axon degeneration remain unknown. We identify endogenous Nmnat2 as a labile axon survival factor whose constant replenishment by anterograde axonal transport is a limiting factor for axon survival. Specific depletion of Nmnat2 is sufficient to induce Wallerian-like degeneration of uninjured axons which endogenous Nmnat1 and Nmnat3 cannot prevent. Nmnat2 is by far the most labile Nmnat isoform and is depleted in distal stumps of injured neurites before Wallerian degeneration begins. Nmnat2 turnover is equally rapid in injured Wld ^S neurites, despite delayed neurite degeneration, showing it is not a consequence of degeneration and also that Wld^S does not stabilize Nmnat2. Depletion of Nmnat2 below a threshold level is necessary for axon degeneration since exogenous Nmnat2 can protect injured neurites when expressed at high enough levels to overcome its short half-life. Furthermore, proteasome inhibition slows both Nmnat2 turnover and neurite degeneration. We conclude that endogenous Nmnat2 prevents spontaneous degeneration of healthy axons and propose that, when present, the more long-lived, functionally related Wld^S protein substitutes for Nmnat2 loss after axon injury. Endogenous Nmnat2 represents an exciting new therapeutic target for axonal disorders.     

Platelet-Mediated Metabolism of the Common Dietary Flavonoid,Quercetin

Background

Flavonoid metabolites remain in blood for periods of time potentially long enough to allow interactions with cellular components of this tissue. It is well-established that flavonoids are metabolised within the intestine and liver into methylated, sulphated and glucuronidated counterparts, which inhibit platelet function.

Methodology/Principal Findings

We demonstrate evidence suggesting platelets which contain metabolic enzymes, as an alternative location for flavonoid metabolism. Quercetin and a plasma metabolite of this compound, 4′-O-methyl quercetin (tamarixetin) were shown to gain access to the cytosolic compartment of platelets, using confocal microscopy. High performance liquid chromatography (HPLC) and mass spectrometry (MS) showed that quercetin was transformed into a compound with a mass identical to tamarixetin, suggesting that the flavonoid was methylated by catechol-O-methyl transferase (COMT) within platelets.

Conclusions/Significance

Platelets potentially mediate a third phase of flavonoid metabolism, which may impact on the regulation of the function of these cells by metabolites of these dietary compounds.     

A Major Histocompatibility Class I Locus Contributes to Multiple Sclerosis Susceptibility Independently from HLA-DRB1*15:01

Background

In Northern European descended populations, genetic susceptibility for multiple sclerosis (MS) is associated with alleles of the human leukocyte antigen (HLA) Class II gene DRB1. Whether other major histocompatibility complex (MHC) genes contribute to MS susceptibility is controversial.

Methodology/Principal Findings

A case control analysis was performed using 958 single nucleotide polymorphisms (SNPs) spanning the MHC assayed in two independent datasets. The discovery dataset consisted of 1,018 cases and 1,795 controls and the replication dataset was composed of 1,343 cases and 1,379 controls. The most significantly MS-associated SNP in the discovery dataset was rs3135391, a Class II SNP known to tag the HLA-DRB1*15:01 allele, the primary MS susceptibility allele in the MHC (O.R. = 3.04, p<1×10⁻⁷⁸). To control for the effects of the HLA-DRB1*15:01 haplotype, case control analysis was performed adjusting for this HLA-DRB1*15:01 tagging SNP. After correction for multiple comparisons (false discovery rate = .05) 52 SNPs in the Class I, II and III regions were significantly associated with MS susceptibility in both datasets using the Cochran Armitage trend test. The discovery and replication datasets were merged and subjects carrying the HLA-DRB1*15:01 tagging SNP were excluded. Association tests showed that 48 of the 52 replicated SNPs retained significant associations with MS susceptibility independently of the HLA-DRB1*15:01 as defined by the tagging SNP. 20 Class I SNPs were associated with MS susceptibility with p-values ≤1×10⁻⁸. The most significantly associated SNP was rs4959039, a SNP in the downstream un-translated region of the non-classical HLA-G gene (Odds ratio 1.59, 95% CI 1.40, 1.81, p = 8.45×10⁻¹³) and is in linkage disequilibrium with several nearby SNPs. Logistic regression modeling showed that this SNP''s contribution to MS susceptibility was independent of the Class II and Class III SNPs identified in this screen.

Conclusions

A MHC Class I locus contributes to MS susceptibility independently of the HLA-DRB1*15:01 haplotype.     

Fingerprinting Diazotroph Communities in the Chesapeake Bay by Using a DNA Macroarray

Investigations of the distribution and diversity of nitrogen-fixing microorganisms in natural environments have often relied on PCR amplification and sequence analysis of a portion of one of the key enzymes in nitrogen fixation, dinitrogenase reductase, encoded by nifH. Recent work has suggested that DNA macroarrays provide semiquantitative fingerprints of diversity within mixtures of nifH amplicons (G. F. Steward, B. D. Jenkins, B. B. Ward, and J. P. Zehr, Appl. Environ. Microbiol. 70:1455-1465, 2004). Here we report the application of macroarrays for a study in the Chesapeake Bay. Samples from different locations in the bay yielded distinct fingerprints. Analysis of replicates and samples from different locations by cluster analysis showed that replicates clustered together, whereas different samples formed distinct clusters. There was a correspondence between the hybridization pattern observed and that predicted from the distribution of sequence types in a corresponding clone library. Some discrepancies between the methods were observed which are likely a result of the high nifH sequence diversity in the Chesapeake Bay and the limited number of sequences represented on this version of the array. Analyses of sequences in the clone library indicate that the Chesapeake Bay harbors unique, phylogenetically diverse diazotrophs. The macroarray hybridization patterns suggest that there are spatially variable communities of diazotrophs, which have been confirmed by quantitative PCR methods (S. M. Short, B. D. Jenkins, and J. P. Zehr, Appl. Environ. Microbiol., in press). The results show that DNA macroarrays have great potential for mapping the spatial and temporal variability of functional gene diversity in the environment.