Vitamin B12 and hepatitis C: Molecular biology and human pathology

Cobalamins are stored in high concentrations in the human liver and thus are available to participate in the regulation of hepatotropic virus functions. We show that cyanocobalamin (vitamin B12) inhibited the H(IV internal ribosome entry site (IRES)-dependent translation of a reporter gene in vitro in a dose-dependent manner without significantly affecting the cap-dependent mechanism. Vitamin B12 failed to inhibit translation by IRES elements from encephalomyocarditis virus (EMCV) or classical swine fever virus (CSFV), We also demonstrate a relationship between the total cobalamin concentration in human sera and HCV viral load (a measure of viral replication in the host), The mean viral load was two orders of magnitude greater when the serum cobalamin concentration was above 200 pM (P < 0.003), suggesting that the total cobalamin concentration in an HCV-infected liver is biologically significant in HCV replication.

Quantitative analysis of the speed and accuracy of tongue movements in dysarthria subsequent to traumatic brain injury using electromagnetic articulography

It has been recognised that in order to study the displacement, timing and co-ordination of articulatory components (i.e., tongue. lips, jaw) in speech production it is desirable to obtain high-resolution movement data on multiple structures inside and outside the vocal tract. Until recently, with the exception of X-ray techniques such as cineradiography, the study 0. speech movements has been hindered by the inaccessibility of the oral cavity during speech. X-ray techniques are generally not used because of unacceptable radiation exposure. The aim of the present study was to demonstrate the use of a new physiological device, the electromagnetic articulograph, for assessing articulatory dysfunction subsequent to traumatic brain injury. The components of the device together with the measuring principle are described and data collected from a single case presented. A 19 year-old male who exhibited dysarthria subsequent to a traumatic brain injury was fitted wit 2 the electromagnetic articulograph (Carstens AG-100) and a kinematic analysis of his tongue movements during production of the lingual consonants it, s, k/ within single syllable words was performed. Examination of kinematic parameters including movemmt trajectories, velocity, and acceleration revealed differences in the speed and accuracy of his tongue movements compared to those produced by a non-neurologically impaired adult male. It was concluded that the articulograph is a useful device for diagnosing speed and accuracy disorders in tongue movements during speech and that the device has potential for incorporation into physiologically based rehabilitation programs as a real-time biofeedback instrument.

Power of the joint segregation analysis method for testing mixed major-gene and polygene inheritance models of quantitative traits

Understanding the genetic architecture of quantitative traits can greatly assist the design of strategies for their manipulation in plant-breeding programs. For a number of traits, genetic variation can be the result of segregation of a few major genes and many polygenes (minor genes). The joint segregation analysis (JSA) is a maximum-likelihood approach for fitting segregation models through the simultaneous use of phenotypic information from multiple generations. Our objective in this paper was to use computer simulation to quantify the power of the JSA method for testing the mixed-inheritance model for quantitative traits when it was applied to the six basic generations: both parents (P-1 and P-2), F-1, F-2, and both backcross generations (B-1 and B-2) derived from crossing the F-1 to each parent. A total of 1968 genetic model-experiment scenarios were considered in the simulation study to quantify the power of the method. Factors that interacted to influence the power of the JSA method to correctly detect genetic models were: (1) whether there were one or two major genes in combination with polygenes, (2) the heritability of the major genes and polygenes, (3) the level of dispersion of the major genes and polygenes between the two parents, and (4) the number of individuals examined in each generation (population size). The greatest levels of power were observed for the genetic models defined with simple inheritance; e.g., the power was greater than 90% for the one major gene model, regardless of the population size and major-gene heritability. Lower levels of power were observed for the genetic models with complex inheritance (major genes and polygenes), low heritability, small population sizes and a large dispersion of favourable genes among the two parents; e.g., the power was less than 5% for the two major-gene model with a heritability value of 0.3 and population sizes of 100 individuals. The JSA methodology was then applied to a previously studied sorghum data-set to investigate the genetic control of the putative drought resistance-trait osmotic adjustment in three crosses. The previous study concluded that there were two major genes segregating for osmotic adjustment in the three crosses. Application of the JSA method resulted in a change in the proposed genetic model. The presence of the two major genes was confirmed with the addition of an unspecified number of polygenes.

Lucerne biology and genetic improvement - an analysis of past activities and future goals in Australia

Breeding methodologies for cultivated lucerne (Medicago sativa L.), an autotetraploid, have changed little over the last 50 years, with reliance on polycross methods and recurrent phenotypic selection. There has been, however, an increase in our understanding of lucerne biology, in particular the genetic relationships between members of the M. sativa complex, as deduced by DNA analysis. Also, the differences in breeding behaviour and vigour of diploids versus autotetraploids, and the underlying genetic causes, are discussed in relation to lucerne improvement. Medicago falcata, a member of the M. sativa complex, has contributed substantially to lucerne improvement in North America, and its diverse genetics would appear to have been under-utilised in Australian programs over the last two decades, despite the reduced need for tolerance to freezing injury in Australian environments. Breeding of lucerne in Australia only commenced on a large scale in 1977, driven by an urgent need to introgress aphid resistance into adapted backgrounds. The release in the early 1980s of lucernes with multiple pest and disease resistance (aphids, Phytophthora, Colletotrichum) had a significant effect on increasing lucerne productivity and persistence in eastern Australia, with yield increases under high disease pressure of up to 300% being recorded over the predominant Australian cultivar, up to 1977, Hunter River. Since that period, irrigated lucerne yields have plateaued, highlighting the need to identify breeding objectives, technologies, and the germplasm that will create new opportunities for increasing performance. This review discusses major goals for lucerne improvement programs in Australia, and provides indications of the germplasm sources and technologies that are likely to deliver the desired outcomes.

Identification of causal relationships among traits related to drought resistance in Stylosanthes scabra using QTL analysis

Previous studies have shown that a negative relationship exists between transpiration efficiency (TE) and carbon isotope discrimination (Delta) and between TE and specific leaf area (SLA) in Stylosanthes scabra, A glasshouse experiment was conducted to confirm these relationships in an F-2 population and to study the causal nature of these relationships through quantitative trait loci (QTL) analysis, One hundred and twenty F-2 genotypes from a cross between two genotypes within S. scabra were used. Three replications for each genotype were maintained through vegetative propagation, Water stress was imposed by maintaining plants at 40% of field capacity for about 45 d. To facilitate QTL analysis, a genetic linkage map consisting of 151 RAPD markers was developed, Results from this study show that Delta was significantly and negatively correlated with TE and biomass production. Similarly, SLA showed significant negative correlation with TE and biomass production, Most of the QTL for TE and Delta were present on linkage groups 5 and 11. Similarly, QTL for SLA, transpiration and biomass productivity traits were clustered on linkage groups 13 and 24, One unlinked marker was also associated with these traits, There were several markers coincident between different traits, At all the coincident QTL, the direction of QTL effects was consistent with phenotypic data, At the coincident markers between TE and Delta, high alleles of TE were associated with low alleles of Delta. Similarly, low alleles of SLA were associated with high alleles of biomass productivity traits and transpiration. At the coincident markers between trans-4-hydroxy-N-methyl proline (MHP) and relative water content (RWC), low alleles of MHP were associated with high alleles of RWC, This study suggests the causal nature of the relationship between TE and Delta. Phenotypic data and QTL, data show that SLA was more closely associated with biomass production than with TE, This study also shows that a cause-effect relationship may exist between SLA and biomass production.

Vascular smooth muscle cell phenotypic modulation in culture is associated with reorganisation of contractile and cytoskeletal proteins

Smooth muscle cells (SMC) exhibit a functional plasticity, modulating from the mature phenotype in which the primary function is contraction, to a less differentiated state with increased capacities for motility, protein synthesis, and proliferation. The present study determined, using Western analysis, double-label immunofluorescence and confocal microscopy, whether changes in phenotypic expression of rabbit aortic SMC in culture could be correlated with alterations in expression and distribution of structural proteins. Contractile state SMC (days 1 and 3 of primary culture) showed distinct sorting of proteins into subcellular domains, consistent with the theory that the SMC structural machinery is compartmentalised within the cell. Proteins specialised for contraction (alpha -SM actin, SM-MHC, and calponin) were highly expressed in these cells and concentrated in the upper central region of the cell. Vimentin was confined to the body of the cell, providing support for the contractile apparatus but not co-localising with it. In line with its role in cell attachment and motility, beta -NM actin was localised to the cell periphery and basal cortex. The dense body protein alpha -actinin was concentrated at the cell periphery, possibly stabilising both contractile and motile apparatus. Vinculin-containing focal adhesions were well developed, indicating the cells' strong adhesion to substrate. In synthetic state SMC (passages 2-3 of culture), there was decreased expression of contractile and adhesion (vinculin) proteins with a concomitant increase in cytoskeletal proteins (beta -non-muscle [NM] actin and vimentin). These quantitative changes in structural proteins were associated with dramatic chan-es in their distribution. The distinct compartmentalisation of structural proteins observed in contractile state SMC was no longer obvious, with proteins more evenly distributed throughout die cytoplasm to accommodate altered cell function. Thus, SMC phenotypic modulation involves not only quantitative changes in contractile and cytoskeletal proteins, but also reorganisation of these proteins. Since the cytoskeleton acts as a spatial regulator of intracellular signalling, reorganisation of the cytoskeleton may lead to realignment of signalling molecules, which, in turn, may mediate the changes in function associated with SMC phenotypic modulation. (C) 2001 Wiley-Liss, Inc.

Parasite infection rather than tactile stimulation is the proximate cause of cleaning behaviour in reef fish

Cleaning behavior is a popular example of non-kin cooperation. However, quantitative support for this is generally sparse and the alternative, that cleaners are parasitic: has also been proposed. Although the behaviour involves some of the most complex and highly developed interspecific communication signals known, the proximate causal factors for why clients Seek cleaners are controversial. However, this information is essential to understanding the evolution of cleaning. I tested whether clients seek cleaners in response to parasite infection or whether clients seek cleaners for tactile stimulation regardless of parasite load. Parasite loads oil client fish were manipulated and clients exposed to cleaner fish and control fish hehind glass. I found that parasitized client fish spent more time than unparasitized fish next to a cleaner fish. In addition; parasitized clients spent more rime next to cleaners than next to control fish whereas unparasitized fish were not attracted to cleaners. This study shows, I believe for the first time, which is somewhat surprising, that parasite infection alone causes clients to seek cleaning by cleaners and provides insight into how this behaviour evolved.

Using MF-PCR to diagnose multiple defects from single cells: implications for PGD

Single cell genetic analysis is generally performed using PCR and FISH. Until recently, FISH has been the method of choice. FISH however is expensive, has significant misdiagnosis rates, can result in interpretation difficulties and is labour intensive making it unsuitable for high throughput processing. Recently fluorescent PCR reliability has increased to levels at or surpassing FISH whilst maintaining low cost. However, PCR accuracy has been a concern due to allelic dropout. Multiplex PCR can now increase accuracy by using multiple markers for each chromosome to firstly provide diagnosis if markers fail and,or secondly confirm diagnosis. We compare a variety of diagnostic methods and demonstrate for the first time a multiplex PCR system providing simultaneous diagnosis and confirmation of the major aneuploidy chromosomes (21, 18, 13) and sex as well as DNA fingerprint in single cells. We also discuss the implications of using PCR for aneuploidy screening in preimplantation genetic diagnosis. (C) 2001 Elsevier Science Ireland Ltd. All rights reserved.

Quantitative evaluation of altered hepatic spaces and membrane transport in fibrotic rat liver

Four animal models were used to quantitatively evaluate hepatic alterations in this study: (1) a carbon tetrachloride control group (phenobarbital treatment only), (2) a CCl4-treated group (phenobarbital with CCl4 treatment), (3) an alcohol-treated group (liquid diet with alcohol treatment), and (4) a pair-fed alcohol control group (liquid diet only). At the end of induction, single-pass perfused livers were used to conduct multiple indicator dilution (MID) studies. Hepatic spaces (vascular space, extravascular albumin space, extravascular sucrose space, and cellular distribution volume) and water hepatocyte permeability/surface area product were estimated from nonlinear regression of outflow concentration versus time profile data. The hepatic extraction ratio of H-3-taurocholate was determined by the nonparametric moments method. Livers were then dissected for histopathologic analyses (e.g., fibrosis index, number of fenestrae). In these 4 models, CCl4-treated rats were found to have the smallest vascular space, extravascular albumin space, H-3-taurocholate extraction, and water hepatocyte permeability/surface area product but the largest extravascular sucrose space and cellular distribution volume. In addition, a linear relationship was found to exist between histopathologic analyses (fibrosis index or number of fenestrae) and hepatic spaces. The hepatic extraction ratio of H-3-taurocholate and water hepatocyte permeability/surface area product also correlated to the severity of fibrosis as defined by the fibrosis index. In conclusion, the multiple indicator dilution data obtained from the in situ perfused rat liver can be directly related to histopathologic analyses.

Convergent maternal care strategies in ungulates and macropods

Mammals show extensive interspecific variation in the form of maternal care. Among ungulates, there is a dichotomy between species in which offspring follow the mother (following strategy) versus species in which offspring remain concealed (hiding strategy). Here we reveal that the same dichotomy exists among macropods (kangaroos, wallabies and allies). We test three traditional adaptive explanations and one new life history hypothesis. and find very similar patterns among both ungulates and macropods. The three traditional explanations that we tested were that a ''following'' strategy is associated with (1) open habitat, (2) large mothers, and (3) gregariousness. Our new life-history hypothesis is that a following strategy'' is associated with delayed weaning, and thus with the slow end of the slow-fast mammalian life-history continuum, because offspring devote resources to locomotion rather than rapid growth. Our comparative test strongly supports the habitat structure hypothesis and provides some support for this new delayed weaning hypothesis for both ungulates and macropods. We propose that sedentary young in closed habitats benefit energetically by having milk brought to them. In open habitats, predation pressure will select against hiding. Followers will suffer slower growth to independence. Taken together, therefore, our results provide the first quantitative evidence that macropods and ungulates are convergent with respect to interspecific variation in maternal care strategy. In both clades, differences between species in the form of parental care are due to a similar interaction between habitat, social behavior, and life history.