Effects of feeding on medicinal leech swimming performance

The locomotor system of sanguivorous leeches is presented with a unique challenge: how to maintain mobility while coping with a >500% increase in body mass during feeding. A meal of this size is likely to disrupt the function of the muscular hydrostat during swimming, reducing speed and increasing predation risks. We quantified the effects of feeding to satiety on swimming kinematics, and the time course of recovery of swimming performance post-feeding in the medicinal leech Hirudo verbana . There was a 5.07 ± 0.04-fold increase in mass during feeding (mean ± sem , n =7). Despite this, leeches were able to swim immediately after feeding, reaching 27% of their pre-feeding speed. Reduced speed was a consequence of a reduction in both swimming cycle frequency and stride length to 69 and 42% of the pre-feeding values, respectively. Recovery of swimming ability was rapid, despite a prolonged increase in body mass. Fifty per cent restoration of swimming speed was achieved in c . 1 h while body mass was still 4.2-fold greater than before feeding. Rapid mass and volume reduction immediately post-feeding, and the properties of the obliquely striated swimming muscles appear to aid recovery of swimming performance. Such features that aid post-feeding recovery of mobility may have been important in the evolution of leech sanguivory.     

Hybrid maize breeding with doubled haploids: V. Selection strategies for testcross performance with variable sizes of crosses and S1 families

In hybrid maize (Zea mays L.) breeding, doubled haploids (DH) are increasingly replacing inbreds developed by recurrent selfing. Doubled haploids may be developed directly from S₀ plants in the parental cross or via S₁ families. In both these breeding schemes, we examined 2 two-stage selecting strategies, i.e., considering or ignoring cross and family structure while selection among and within parental crosses and S₁ families. We examined the optimum allocation of resources to maximize the selection gain ΔG and the probability P(q) of identifying the q% best genotypes. Our specific objectives were to (1) determine the optimum number and size of crosses and S₁ families, as well as the optimum number of test environments and (2) identify the superior selection strategy. Selection was based on the evaluation of testcross progenies of (1) DH lines in both stages (DHTC) and (2) S₁ families in the first stage and of DH lines within S₁ families in the second stage (S₁TC-DHTC) with uniform and variable sizes of crosses and S₁ families. We developed and employed simulation programs for selection with variable sizes of crosses and S₁ families within crosses. The breeding schemes and selection strategies showed similar relative efficiency for both optimization criteria ΔG and P (0.1%). As compared with DHTC, S₁TC-DHTC had larger ΔG and P (0.1%), but a higher standard deviation of ΔG. The superiority of S₁TC-DHTC was increased when the selection was done among all DH lines ignoring their cross and family structure and using variable sizes of crosses and S₁ families. In DHTC, the best selection strategy was to ignore cross structures and use uniform size of crosses.     

High congruency of QTL positions for heterosis of grain yield in three crosses of maize

The genetic basis of heterosis in maize has been investigated in a number of studies but results have not been conclusive. Here, we compare quantitative trait loci (QTL) mapping results for grain yield, grain moisture, and plant height from three populations derived from crosses of the heterotic pattern Iowa Stiff Stalk Synthetic × Lancaster Sure Crop, investigated with the Design III, and analyzed with advanced statistical methods specifically developed to examine the genetic basis of mid-parent heterosis (MPH). In two populations, QTL analyses were conducted with a joint fit of linear transformations Z ₁ (trait mean across pairs of backcross progenies) and Z ₂ (half the trait difference between pairs of backcross progenies) to estimate augmented additive and augmented dominance effects of each QTL, as well as their ratio. QTL results for the third population were obtained from the literature. For Z ₂ of grain yield, congruency of QTL positions was high across populations, and a large proportion of the genetic variance (~70%) was accounted for by QTL. This was not the case for Z ₁ or the other two traits. Further, almost all congruent grain yield QTL were located in the same or an adjacent bin encompassing the centromere. We conclude that different alleles have been fixed in each heterotic pool, which in combination with allele(s) from the opposite heterotic pool lead to high MPH for grain yield. Their positive interactions very likely form the base line for the superior performance of the heterotic pattern under study.     

Proteome of conidial surface associated proteins of Aspergillus fumigatus reflecting potential vaccine candidates and allergens

Aspergillus fumigatus is a mold causing most of the invasive fungal lung infections in the immunocompromised host. In addition, the species is the causative agent of certain allergic diseases. Both in invasive and in allergic diseases, the conidial surface mediates the first contact with the human immune system. Thus, conidial surface proteins may be reasonable vaccine candidates as well as important allergens. To broaden the list of those antigens, intact viable Aspergillus conidia were extracted with mild alkaline buffer at pH 8.5 in the presence of a 1,3-beta-glucanase. The proteome of this fraction was separated by two- dimensional gel electrophoresis (2-DE) and analyzed by liquid chromatography coupled with tandem mass spectrometry. Altogether 26 different A. fumigatus proteins were identified, twelve of which contain a signal for secretion. Among these were the known major conidial surface protein rodlet A, one acid protease PEP2, one lipase, a putative disulfide isomerase and a putative fructose-1,6-biphosphatase. The known allergen Aspf 3 was identified among the proteins without a signal for secretion. On the basis of the recently annotated A. fumigatus genome (Nature 2005, 438, 1151-1156), proteome analysis is now a powerful tool to confirm expression of hypothetical proteins and, thereby to identify additional vaccine candidates and possible new allergens of this important fungal pathogen.     

A new two-color Fab labeling method for autoantigen protein microarrays

Antigen microarrays hold great promise for profiling the humoral immune response in the settings of autoimmunity, allergy and cancer. This approach involves immobilizing antigens on a slide surface and then exposing the array to biological fluids containing immunoglobulins. Although these arrays have proven extremely useful as research tools, they suffer from several sources of variability. To address these issues, we have developed a new two-color Fab labeling method that allows two samples to be applied simultaneously to the same array. This straightforward labeling approach improves reproducibility and reliably detects changes in autoantibody concentrations. Using this technique we profiled serum from a mouse model of systemic lupus erythematosus (SLE) and detected both expected and previously unrecognized reactivities. The improved labeling and detection method described here overcomes several problems that have hindered antigen microarrays and should facilitate translation to the clinical setting.     

High base pair opening rates in tracts of GC base pairs

Sequence-dependent structural features of the DNA double helix have a strong influence on the base pair opening dynamics. Here we report a detailed study of the kinetics of base pair breathing in tracts of GC base pairs in DNA duplexes derived from 1H NMR measurements of the imino proton exchange rates upon titration with the exchange catalyst ammonia. In the limit of infinite exchange catalyst concentration, the exchange times of the guanine imino protons of the GC tracts extrapolate to much shorter base pair lifetimes than commonly observed for isolated GC base pairs. The base pair lifetimes in the GC tracts are below 5 ms for almost all of the base pairs. The unusually rapid base pair opening dynamics of GC tracts are in striking contrast to the behavior of AT tracts, where very long base pair lifetimes are observed. The implication of these findings for the structural principles governing spontaneous helix opening as well as the DNA-binding specificity of the cytosine-5-methyltransferases, where flipping of the cytosine base has been observed, are discussed.     

Quantitative trait locus mapping based on resampling in a vast maize testcross experiment and its relevance to quantitative genetics for complex traits

From simulation studies it is known that the allocation of experimental resources has a crucial effect on power of QTL detection as well as on accuracy and precision of QTL estimates. In this study, we used a very large experimental data set composed of 976 F(5) maize testcross progenies evaluated in 19 environments and cross-validation to assess the effect of sample size (N), number of test environments (E), and significance threshold on the number of detected QTL, the proportion of the genotypic variance explained by them, and the corresponding bias of estimates for grain yield, grain moisture, and plant height. In addition, we used computer simulations to compare the usefulness of two cross-validation schemes for obtaining unbiased estimates of QTL effects. The maximum, validated genotypic variance explained by QTL in this study was 52.3% for grain moisture despite the large number of detected QTL, thus confirming the infinitesimal model of quantitative genetics. In both simulated and experimental data, the effect of sample size on power of QTL detection as well as on accuracy and precision of QTL estimates was large. The number of detected QTL and the proportion of genotypic variance explained by QTL generally increased more with increasing N than with increasing E. The average bias of QTL estimates and its range were reduced by increasing N and E. Cross-validation performed well with respect to yielding asymptotically unbiased estimates of the genotypic variance explained by QTL. On the basis of our findings, recommendations for planning of QTL mapping experiments and allocation of experimental resources are given.     

Development of a periodontium-approximated specimen holder for attrition studies of telescopic crowns]

To enable attritional studies on telescopic crowns, a specimen holder has been constructed that is intended to simulate, in zero approximation, the biomechanical properties of abutments bearing telescopic crowns. The specimen holder consists essentially of a system with two metal "concertinas" with different elasticity constants reflecting the different biomechanical parameters for periodontium and bone. The biomechanical processes in the periodontium are too complex to be simulated accurately in vitro. Nevertheless, the specimen holder described here enables the forces acting during insertion and removal of a partial denture to be accommodated elastically in the angular and axial directions, to simulate the possible degrees of freedom of the abutments in situ. In this way, the unavoidable errors of specimen fixation are rendered tolerable, so that local preferential wear is decisively reduced.     

Protein microarrays guide tolerizing DNA vaccine treatment of autoimmune encephalomyelitis

The diversity of autoimmune responses poses a formidable challenge to the development of antigen-specific tolerizing therapy. We developed 'myelin proteome' microarrays to profile the evolution of autoantibody responses in experimental autoimmune encephalomyelitis (EAE), a model for multiple sclerosis (MS). Increased diversity of autoantibody responses in acute EAE predicted a more severe clinical course. Chronic EAE was associated with previously undescribed extensive intra- and intermolecular epitope spreading of autoreactive B-cell responses. Array analysis of autoantigens targeted in acute EAE was used to guide the choice of autoantigen cDNAs to be incorporated into expression plasmids so as to generate tolerizing vaccines. Tolerizing DNA vaccines encoding a greater number of array-determined myelin targets proved superior in treating established EAE and reduced epitope spreading of autoreactive B-cell responses. Proteomic monitoring of autoantibody responses provides a useful approach to monitor autoimmune disease and to develop and tailor disease- and patient-specific tolerizing DNA vaccines.