The probabilistic economic injury level: incorporating uncertainty into pest management decision-making

Despite the recognition that the economic injury level (EIL) is determined by dynamic biological and economic parameters, which can be highly variable and uncertain, there has been little effort to quantify uncertainty and to use estimates of uncertainty in the determination of EILs. In this paper, we define the probabilistic EIL (PEIL) and develop PEILs for two insect pest scenarios: alfalfa weevil larvae, Hypera postica (Gyllenhal), on early bud-stage alfalfa, and bean leaf beetle adults, Cerotoma trifurcata (Forster), on V1-stage soybean. The PEIL is an EIL that reflects its probability of occurrence. The probability of occurrence is determined by incorporating the uncertainty associated with the input variables used to calculate the EIL. We used Monte Carlo simulation, a random sampling technique in which each input variable in the model was sampled repeatedly from a range of possible values based on probability distributions. Each input variable's probability distribution was sampled such that the distribution's shape was reproduced. Then, the variability for each input was propagated into the output of the model so that the model output reflected the probability of values that could occur. This represents the first use of the Monte Carlo technique to determine EILs.     

Measurements of photosynthesis and respiration in plants

Methods for measuring the rates of photosynthesis and respiration in plants are reviewed. Closed systems that involve manometric techniques, ¹⁴CO₂ fixation, O₂ electrodes and other methods for measuring dissolved and gas phase O₂ are described. These methods typically provide time-integrated rate measurements, and limitations to their use are discussed. Open gas exchange systems that use infra-red CO₂ gas analysers and differential O₂ analysers for measuring instantaneous rates of CO₂ and O₂ exchange are described. Important features of the analysers, design features of gas exchange systems, and sources of potential error are considered. The analysis of chlorophyll fluorescence parameters for estimating the quantum yield for O₂ evolution and CO₂ fixation is described in relation to new fluorescence imaging systems for large scale screening of photosynthetic phenotypes, and the microimaging of individual chloroplasts.     

Cryopreservation of umbilical cord blood: 1. Osmotically inactive volume,hydraulic conductivity and permeability of CD34(+) cells to dimethyl sulphoxide

Umbilical cord blood (UCB) is an accepted treatment for the reconstitution of bone marrow function following myeloablative treatment predominantly in children and juveniles. Current cryopreservation protocols use methods established for bone marrow and peripheral blood progenitors cells that have largely been developed empirically. Such protocols can result in losses of up to 50% of the nucleated cell population: losses unacceptable for cord blood. The design of optimal cryopreservation regimes requires the development of addition and elution protocols for the chosen cryoprotectant; protocols that minimise damaging osmotic transients. The biophysical parameters necessary to model the addition and elution of dimethyl sulphoxide to and from cord blood CD34(+) cells have been established. An electronic particle counting method was used to establish the volumetric response of CD34(+) cells to changes in osmolality of the suspending medium. The non-osmotic volume of the cell was 0.27 of the cells isotonic volume. The permeation kinetics of CD34(+) cells to water and dimethyl sulphoxide were investigated at two temperatures, +1.5 and +20 degrees C. Values for the hydraulic conductivity were 3.2 x 10(-8) and 2.8 x 10(-7)cm/atm/s, respectively. Values for the permeability of dimethyl sulphoxide at these temperatures were 4.2 x 10(-7) and 7.4 x 10(-6)cm/s, respectively. Clonogenic assays indicated that the ability of CD34(+) cells to grow and differentiate was significantly impaired outside the limits 0.6-4x isotonic. Based on the Boyle van't Hoff plot, the tolerable limits for cell volume excursion were therefore 45-140% of isotonic volume. The addition and elution of cryoprotectant was modelled using a two-parameter model. Current protocols for the addition of cryoprotectant based on exposure at +4 degrees C would require additional time for complete equilibration of the cryoprotectant. During the elution phase current protocols are likely to cause CD34(+) cells to exceed tolerable limits. The addition of a short holding period during elution reduces the likelihood of this occurring.     

The single species hypothesis: truly dead and pushing up bushes, or still twitching and ripe for resuscitation?

Frank Livingstone proclaims himself to be the last living proponent of the single species hypothesis. In sharp contrast, a species-rich, bushy phylogeny is favored by most human paleontologists. Is Livingstone's proclamation merely contrarian posturing, or does closer inspection warrant reconsideration of just how speciose the hominin lineage is? The high-speciation perspective draws on evidence of speciosity in the Cercopithecoidea and punctuated equilibria theory for support. If blue monkeys and redtail monkeys are indistinguishable skeletally, this reasoning goes, or if red colobus and black and white colobus are likewise indistinguishable, should we not expect that there are more species of hominin than is apparent from skeletal evidence alone? A contrarian perspective notes that not all monkey taxa are speciose. Importantly, two broadly distributed, partly terrestrial monkeys have not speciated at all: vervets and baboons. Nor are monkeys the first choice as a hominin speciation model. If expectations of species numbers are based on the Hominoidea, a taxon more closely related to hominins, more similar in body size, and found in more hominin-like habitats than monkeys, a single-species perspective is more appealing. No great ape genus has even two sympatric species. Moreover, despite a separation of 1.6 Ma, West African chimpanzees have not speciated from Pt. troglodytes nor Pt. schweinfurthii. It is notable that no two contemporaneous species of hominin were separated by significantly more than this interval. A biological--as opposed to an ecological or geographical--species definition would place all hominins in a single, phenotypically diverse species. Since divergence from the chimpanzee, "species" distinctness in hominins may have been maintained by temporary allopatry and centripetal niche separation. The hominin lineage may have evolved as a single, phenotypically diverse, reticulately evolving species.     

Crystal structures of the BtuF periplasmic-binding protein for vitamin B12 suggest a functionally important reduction in protein mobility upon ligand binding

BtuF is the periplasmic binding protein (PBP) for the vitamin B12 transporter BtuCD, a member of the ATP-binding cassette (ABC) transporter superfamily of transmembrane pumps. We have determined crystal structures of Escherichia coli BtuF in the apo state at 3.0 A resolution and with vitamin B12 bound at 2.0 A resolution. The structure of BtuF is similar to that of the FhuD and TroA PBPs and is composed of two alpha/beta domains linked by a rigid alpha-helix. B12 is bound in the "base-on" or vitamin conformation in a wide acidic cleft located between these domains. The C-terminal domain shares structural homology to a B12-binding domain found in a variety of enzymes. The same surface of this domain interacts with opposite surfaces of B12 when comparing ligand-bound structures of BtuF and the homologous enzymes, a change that is probably caused by the obstruction of the face that typically interacts with this domain by the base-on conformation of vitamin B12 bound to BtuF. There is no apparent pseudo-symmetry in the surface properties of the BtuF domains flanking its B12 binding site even though the presumed transport site in the previously reported crystal structure of BtuCD is located in an intersubunit interface with 2-fold symmetry. Unwinding of an alpha-helix in the C-terminal domain of BtuF appears to be part of conformational change involving a general increase in the mobility of this domain in the apo structure compared with the B12-bound structure. As this helix is located on the surface likely to interact with BtuC, unwinding of the helix upon binding to BtuC could play a role in triggering release of B12 into the transport cavity. Furthermore, the high mobility of this domain in free BtuF could provide an entropic driving force for the subsequent release of BtuF required to complete the transport cycle.     

The long form of CDK2 arises via alternative splicing and forms an active protein kinase with cyclins A and E

We have reinvestigated the long form of cyclin-dependent kinase (CDK)2 that is expressed in many rodent cells. We show that the mRNA encoding CDK2L arises by alternative splicing and that the encoded protein can bind to, and be activated by, cyclins A and E. The complex of CDK2L with cyclin A has about half the specific activity of the equivalent CDK2-cyclin A complex. Also, CDK2L--cyclin A is inhibited to the same extent and by the same concentrations of p21(CIP1) as CDK2--cyclin A. The nucleotide sequences of intron V in the human and murine CDK2 genes, where the sequences encoding the 48-residue insert in CDK2L are located, show very high conservation in the position of the alternatively spliced exon and its surroundings. Despite this, we were not able to detect significant expression of CDK2L in human cell lines, although a low level is expressed in COS-1 cells from monkeys.     

Protein solubility and folding monitored in vivo by structural complementation of a genetic marker protein

Protein misfolding is the basis of a number of human diseases and presents an obstacle to the production of soluble recombinant proteins. We present a general method to assess the solubility and folding of proteins in vivo. The basis of this assay is structural complementation between the alpha- and omega- fragments of beta-galactosidase (beta-gal). Fusions of the alpha-fragment to the C terminus of target proteins with widely varying in vivo folding yield and/or solubility levels, including the Alzheimer's amyloid beta (A beta) peptide and a non-amyloidogenic mutant thereof, reveal an unambiguous correlation between beta-gal activity and the solubility/folding of the target. Thus, structural complementation provides a means of monitoring protein solubility/misfolding in vivo, and should find utility in the screening for compounds that influence the pathological consequences of these processes.     

The molecular dynamics of pain control

Pain is necessary for survival, but persistent pain can result in anxiety, depression and a reduction in the quality of life. The discriminative and affective qualities of pain are both thought to be regulated in an activity-dependent fashion. Recent studies have identified cells and molecules that regulate pain sensitivity and the parallel pathways that distribute nociceptive information to limbic or sensory areas of the forebrain. Here, we emphasize the cellular and neurobiological consequences of pain, especially those that are involved in the generation and maintenance of chronic pain. These new insights into pain processing will significantly alter our approach to pain control and the development of new analgesics.     

Central nervous system in cerebral malaria: 'Innocent bystander' or active participant in the induction of immunopathology?

Cerebral malaria (CM) is a major life-threatening complication of Plasmodium falciparum infection in humans, responsible for up to 2 million deaths annually. The mechanisms underlying the fatal cerebral complications are still not fully understood. Many theories exist on the aetiology of human CM. The sequestration hypo-thesis suggests that adherence of parasitized erythrocytes to the cerebral vasculature leads to obstruction of the microcirculation, anoxia or metabolic disturbances affecting brain function, resulting in coma. This mechanism alone seems insufficient to explain all the known features of CM. In this review we focus on another major school of thought, that CM is the result of an over-vigorous immune response originally evolved for the protection of the host. Evidence in support of this second hypothesis comes from studies in murine malaria models in which T cells, monocytes, adhesion molecules and cytokines, have been implicated in the development of the cerebral complications. Recent studies of human CM also indicate a role for the immune system in the neurological complications. However, it is likely that multiple mechanisms are involved in the induction of cerebral complications and both the presence of parasitized erythrocytes in the central nervous system (CNS) and immunopathological processes contribute to the pathogenesis of CM. Most studies examining immunopathological responses in CM have focused on reactions occurring primarily in the systemic circulation. However, these also do not fully account for the development of cerebral complications in CM. In this review we summarize results from human and mouse studies that demonstrate morphological and functional changes in the resident glial cells of the CNS. The degree of immune activation and degeneration of glial cells was shown to reflect the extent of neurological complications in murine cerebral malaria. From these results we highlight the need to consider the potentially important contribution within the CNS of glia and their secreted products, such as cytokines, in the development of human CM.     

The intracellular oxidation of 2',7'-dichlorofluorescin in murine T lymphocytes

Intracellular reactive oxygen species (ROS) production by activated murine T lymphocytes was investigated by analyzing intracellular dichlorofluorescin (DCFH(2)) oxidation in lymph node cells (LNC). An increase in DCFH(2) oxidation in LNC induced by phorbol myristate acetate (PMA) was detected by flow cytometry. It was confirmed that this increase was present in Thy1(+) LNC. We examined the contribution to intracellular DCFH(2) oxidation of ROS released by leukocytes other than T cells present in the LNC suspension. Superoxide dismutase, catalase, and glutathione/glutathione peroxidase inhibited the PMA-induced increase in intracellular DCFH(2) oxidation. Furthermore, PMA failed to elicit DCFH(2) oxidation in LNC isolated from mice lacking a functional NADPH oxidase (gp91(phox) gene knockout mice), but this response could be restored in these cells by the addition of T cell-depleted LNC from wild-type litter mates. This study highlights the necessity for caution in using the DCFH(2) assay to demonstrate specific intracellular ROS production in heterogeneous cell populations. It also suggests that cells other than T cells in lymph node populations may, through production of ROS, influence the intracellular redox state of T lymphocytes.