Influenza antibody breadth and effector functions are immune correlates from acquisition of pandemic infection of children.

Cross-reactive antibodies with Fc receptor (FcR) effector functions may mitigate pandemic virus impact in the absence of neutralizing antibodies. In this exploratory study, we use serum from a randomized placebo-controlled trial of seasonal trivalent influenza vaccination in children (NCT00792051) conducted at the onset of the 2009 H1N1 pandemic (pH1N1) and monitored for infection. We found that seasonal vaccination increases pH1N1 specific antibodies and FcR effector functions. Furthermore, prospective baseline antibody profiles after seasonal vaccination, prior to pH1N1 infection, show that unvaccinated uninfected children have elevated ADCC effector function, FcγR3a and FcγR2a binding antibodies to multiple pH1N1 proteins, past seasonal and avian (H5, H7 and H9) strains. Whereas, children that became pH1N1 infected after seasonal vaccination have antibodies focussed to seasonal strains without FcR functions, and greater aggregated HA-specific profiles for IgM and IgG3. Modeling to predict infection susceptibility, ranked baseline hemagglutination antibody inhibition as the highest contributor to lack of pH1N1 infection, in combination with features that include pH1-IgG1, H1-stem responses and FcR binding to seasonal vaccine and pH1 proteins. Thus, seasonal vaccination can have benefits against pandemic influenza viruses, and some children already have broadly reactive antibodies with Fc potential without vaccination and may be considered 'elite influenza controllers'.

Tissue-resident glial cells associate with tumoral vasculature and promote cancer progression.

Cancer cells are embedded within the tissue and interact dynamically with its components during cancer progression. Understanding the contribution of cellular components within the tumor microenvironment is crucial for the success of therapeutic applications. Here, we reveal the presence of perivascular GFAP+/Plp1+ cells within the tumor microenvironment. Using in vivo inducible Cre/loxP mediated systems, we demonstrated that these cells derive from tissue-resident Schwann cells. Genetic ablation of endogenous Schwann cells slowed down tumor growth and angiogenesis. Schwann cell-specific depletion also induced a boost in the immune surveillance by increasing tumor-infiltrating anti-tumor lymphocytes, while reducing immune-suppressor cells. In humans, a retrospective in silico analysis of tumor biopsies revealed that increased expression of Schwann cell-related genes within melanoma was associated with improved survival. Collectively, our study suggests that Schwann cells regulate tumor progression, indicating that manipulation of Schwann cells may provide a valuable tool to improve cancer patients' outcomes.

A Quantitative Approach to Unravel the Role of Host Genetics in IgG-FcγR Complex Formation After Vaccination.

Fc-mediated immune functions have been correlated with protection in the RV144 HIV vaccine trial and are important for immunity to a range of pathogens. IgG antibodies (Abs) that form complexes with Fc receptors (FcRs) on innate immune cells can activate Fc-mediated immune functions. Genetic variation in both IgGs and FcRs have the capacity to alter IgG-FcR complex formation via changes in binding affinity and concentration. A growing challenge lies in unraveling the importance of multiple variations, especially in the context of vaccine trials that are conducted in homogenous genetic populations. Here we use an ordinary differential equation model to quantitatively assess how IgG1 allotypes and FcγR polymorphisms influence IgG-FcγRIIIa complex formation in vaccine-relevant settings. Using data from the RV144 HIV vaccine trial, we map the landscape of IgG-FcγRIIIa complex formation predicted post-vaccination for three different IgG1 allotypes and two different FcγRIIIa polymorphisms. Overall, the model illustrates how specific vaccine interventions could be applied to maximize IgG-FcγRIIIa complex formation in different genetic backgrounds. Individuals with the G1m1,17 and G1m1,3 allotypes were predicted to be more responsive to vaccine adjuvant strategies that increase antibody FcγRIIIa affinity (e.g. glycosylation modifications), compared to the G1m-1,3 allotype which was predicted to be more responsive to vaccine boosting regimens that increase IgG1 antibody titers (concentration). Finally, simulations in mixed-allotype populations suggest that the benefit of boosting IgG1 concentration versus IgG1 affinity may be dependent upon the presence of the G1m-1,3 allotype. Overall this work provides a quantitative tool for rationally improving Fc-mediated functions after vaccination that may be important for assessing vaccine trial results in the context of under-represented genetic populations.

Serological and cellular inflammatory signatures in end-stage kidney disease and latent tuberculosis.

OBJECTIVES: Tuberculosis comorbidity with chronic diseases including diabetes, HIV and chronic kidney disease is of rising concern. In particular, latent tuberculosis infection (LTBI) comorbidity with end-stage kidney disease (ESKD) is associated with up to 52.5-fold increased risk of TB reactivation to active tuberculosis infection (ATBI). The immunological mechanisms driving this significant rise in TB reactivation are poorly understood. To contribute to this understanding, we performed a comprehensive assessment of soluble and cellular immune features amongst a unique cohort of patients comorbid with ESKD and LTBI. METHODS: We assessed the plasma and cellular immune profiles from patients with and without ESKD and/or LTBI (N = 40). We characterised antibody glycosylation, serum complement and cytokine levels. We also assessed classical and non-classical monocytes and T cells with flow cytometry. Using a systems-based approach, we identified key immunological features that discriminate between the different disease states. RESULTS: Individuals with ESKD exhibited a highly inflammatory plasma profile and an activated cellular state compared with those without ESKD, including higher levels of inflammatory antibody Fc glycosylation structures and activated CX3CR1+ monocytes that correlate with increased inflammatory plasma cytokines. Similar elevated inflammatory signatures were also observed in ESKD+/LTBI+ compared with ESKD-/LTBI+, suggesting that ESKD induces an overwhelming inflammatory immune state. In contrast, no significant inflammatory differences were observed when comparing LTBI+ and LTBI- individuals. CONCLUSION: Our study highlights the highly inflammatory state induced by ESKD. We hypothesise that this inflammatory state could contribute to the increased risk of TB reactivation in ESKD patients.

A systems approach to elucidate personalized mechanistic complexities of antibody-Fc receptor activation post-vaccination.

Immunoglobulin G (IgG) antibodies that activate Fc-mediated immune functions have been correlated with vaccine efficacy, but it is difficult to unravel the relative roles of multiple IgG and Fc receptor (FcR) features that have the capacity to influence IgG-FcR complex formation but vary on a personalized basis. Here, we develop an ordinary differential-equation model to determine how personalized variability in IgG subclass concentrations and binding affinities influence IgG-FcγRIIIa complex formation and validate it with samples from the HIV RV144 vaccine trial. The model identifies individuals who are sensitive, insensitive, or negatively affected by increases in HIV-specific IgG1, which is validated with the addition of HIV-specific IgG1 monoclonal antibodies to vaccine samples. IgG1 affinity to FcγRIIIa is also prioritized as the most influential parameter for dictating activation broadly across a population. Overall, this work presents a quantitative tool for evaluating personalized differences underlying FcR activation, which is relevant to ongoing efforts to improve vaccine efficacy.

An Inflammatory Story: Antibodies in Tuberculosis Comorbidities.

Mycobacterium tuberculosis (Mtb) resides in a quarter of the world's population and is the causative agent for tuberculosis (TB), the most common infectious reason of death in humans today. Although cellular immunity has been firmly established in the control of Mtb, there is growing evidence that antibodies may also modulate the infection. More specifically, certain antibody features are associated with inflammation and are divergent in different states of human infection and disease. Importantly, TB impacts not just the healthy but also those with chronic conditions. While HIV represents the quintessential comorbid condition for TB, recent epidemiological evidence shows that additional chronic conditions such as diabetes and kidney disease are rising. In fact, the prevalence of diabetes as a comorbid TB condition is now higher than that of HIV. These chronic diseases are themselves independently associated with pro-inflammatory immune states that encompass antibody profiles. This review discusses isotypes, subclasses, post-translational modifications and Fc-mediated functions of antibodies in TB infection and in the comorbid chronic conditions of HIV, diabetes, and kidney diseases. We propose that inflammatory antibody profiles, which are a marker of active TB, may be an important biomarker for detection of TB disease progression within comorbid individuals. We highlight the need for future studies to determine which inflammatory antibody profiles are the consequences of comorbidities and which may potentially contribute to TB reactivation.

Reproductive traits and evolutionary divergence between Mediterranean crops and their wild relatives.

Changes in reproductive traits associated with domestication critically determine the evolutionary divergence between crops and their wild relatives, as well as the potential of crop plants to become feral. In this review, we examine the genetic mechanisms of plant domestication and the different types of selection involved, and describe the particularities of domestication of Mediterranean field crops with regard to their reproductive traits, showing illustrative examples. We also explore gene flow patterns between Mediterranean field crops and their wild relatives, along with their ecological, evolutionary and economic implications. Domestication entails multiple selective processes, including direct selection, environmental adaptation and developmental constraints. In contrast to clonal propagation in perennials, sexual reproduction and seed propagation in annuals and biennials have led to a distinct pathway of evolution of reproductive traits. Thus, the initial domestication and further breeding of Mediterranean field crops has brought about changes in reproductive traits, such as higher mean values and variance of seed and fruit sizes, reduced fruit and seed toxicity, non-shattering seeds and loss of seed dormancy. Evolution under domestication is not a linear process, and bi-directional gene flow between wild and crop taxa is a frequent phenomenon. Thus, hybridisation and introgression have played a very important role in determining the genetics of current cultivars. In turn, gene flow from crops to wild relatives can lead to introgression of crop genes into wild populations and potentially alter the characteristics of natural communities. In conclusion, plant evolution under domestication has not only changed the reproductive biology of cultivated taxa, its effects are multifaceted and have implications beyond agriculture.

Dimeric Fcγ Receptor Enzyme-Linked Immunosorbent Assay To Study HIV-Specific Antibodies: A New Look into Breadth of Fcγ Receptor Antibodies Induced by the RV144 Vaccine Trial.

Ab-dependent cellular cytotoxicity (ADCC) responses are of growing interest in the HIV vaccine field but current cell-based assays are usually difficult to reproduce across laboratories. We developed an ELISA and multiplex assay to model the cross-linking of Fcγ receptors (FcγR) by Abs, which is required to initiate an ADCC response. Our FcγR dimer ELISA readily detected Abs in samples from two separate cohorts of the partially efficacious Thai RV144 HIV vaccine efficacy trial. The FcγR dimer-binding Abs induced by the RV144 regimen correlated well with a functional measure of ADCC as well as IgG subclasses. The high-throughput multiplex assay allowed us to simultaneously measure FcγR dimer-binding Abs to 32 different HIV Ags, providing a measure of the breadth of FcγR-binding Abs induced by the RV144 trial. FcγR-binding Abs specific to V regions 1 and 2 were strongly associated with increased breadth of recognition of different Env proteins, suggesting anti-V regions 1 and 2 Abs may be a marker of ADCC breadth. This FcγR dimer provides an important tool for the further analysis and refinement of ADCC-inducing HIV and other antiviral vaccine regimens.

Effect of Combination Antiretroviral Therapy on HIV-1-specific Antibody-Dependent Cellular Cytotoxicity Responses in Subtype B- and Subtype C-Infected Cohorts.

BACKGROUND: There is growing interest in immune therapies to clear the latent HIV-1 after combination antiretroviral therapy (cART). There is limited information on the effect of cART on antibody-dependent cellular cytotoxicity (ADCC), and no studies have directly compared ADCC in HIV-1 subtype B- and subtype C-infected subjects. The effect of improving immunocompetence on ADCC to influenza also remains unexplored. METHODS: The effect of cART on HIV-1- and influenza-specific ADCC was analyzed in 2 cohorts (39 subtype B- and 47 subtype C-infected subjects) before and after 2 years of cART. ADCC analyses included an enzyme-linked immunosorbent assay-based dimeric recombinant soluble (rs) FcγRIIIa-binding assay, antibody-dependent natural killer cell activation assay, and ADCC-mediated killing assays. RESULTS: HIV-1 subtype B and C Env-specific antibody binding to dimeric rsFcγRIIIa were reduced in subtypes B- and C-infected cohorts after 2 years of cART (both P < 0.05). Reduced ADCC-mediated killing of target cells expressing subtype B Env in the subtype B-infected cohort (P = 0.003) was observed after 96 weeks of cART, but not of subtype C Env in the subtype C-infected cohort. A greater reduction in ADCC was detected in subjects with baseline CD4 counts >300 cells/µL (P < 0.05). The resolving immunodeficiency after 96 weeks of cART resulted in improved HA-specific ADCC to 6 strains of influenza (all P < 0.01). CONCLUSIONS: cART results in HIV-1 antigen loss and reductions in HIV-1 Env-specific antibodies with Fc functionality in both subtype B- and C-infected subjects, particularly in immunocompetent subjects. Simultaneously, cART improves ADCC to diverse strains of influenza, suggesting reduction in influenza disease after cART.

Antibody Functional Assays as Measures of Fc Receptor-Mediated Immunity to HIV - New Technologies and their Impact on the HIV Vaccine Field.

BACKGROUND: There is now intense interest in the role of HIV-specific antibodies and the engagement of FcγR functions in the control and prevention of HIV infection. The analyses of the RV144 vaccine trial, natural progression cohorts, and macaque models all point to a role for Fc-dependent effector functions, such as cytotoxicity (ADCC) or phagocytosis (ADCP), in the control of HIV. However, reliable assays that can be reproducibly used across different laboratories to measure Fcdependent functions, such as antibody dependent cellular cytotoxicity (ADCC) are limited. METHOD: This brief review highlights the importance of Fc properties for immunity to HIV, particularly via FcγR diversity and function. We discuss assays used to study FcR mediated functions of HIV-specific Ab, including our recently developed novel cell-free ELISA using homo-dimeric FcγR ectodomains to detect functionally relevant viral antigen-specific antibodies. RESULTS: The binding of these dimeric FcγR ectodomains, to closely spaced pairs of IgG Fc, mimics the engagement and cross-linking of Fc receptors by IgG opsonized virions or infected cells as the essential prerequisite to the induction of Ab-dependent effector functions. The dimeric FcγR ELISA reliably correlates with ADCC in patient responses to influenza. The assay is amenable to high throughput and could be standardized across laboratories. CONCLUSION: We propose the assay has broader implications for the evaluation of the quality of antibody responses in viral infections and for the rapid evaluation of responses in vaccine development campaigns for HIV and other viral infections.

Growing larger with domestication: a matter of physiology, morphology or allocation?

Domestication might affect plant size. We investigated whether herbaceous crops are larger than their wild progenitors, and the traits that influence size variation. We grew six crop plants and their wild progenitors under common garden conditions. We measured the aboveground biomass gain by individual plants during the vegetative stage. We then tested whether photosynthesis rate, biomass allocation to leaves, leaf size and specific leaf area (SLA) accounted for variations in whole-plant photosynthesis, and ultimately in aboveground biomass. Despite variations among crops, domestication generally increased the aboveground biomass (average effect +1.38, Cohen's d effect size). Domesticated plants invested less in leaves and more in stems than their wild progenitors. Photosynthesis rates remained similar after domestication. Variations in whole-plant C gains could not be explained by changes in leaf photosynthesis. Leaves were larger after domestication, which provided the main contribution to increases in leaf area per plant and plant-level C gain, and ultimately to larger aboveground biomass. In general, cultivated plants have become larger since domestication. In our six crops, this occurred despite lower investment in leaves, comparable leaf-level photosynthesis and similar biomass costs of leaf area (i.e. SLA) than their wild progenitors. Increased leaf size was the main driver of increases in aboveground size. Thus, we suggest that large seeds, which are also typical of crops, might produce individuals with larger organs (i.e. leaves) via cascading effects throughout ontogeny. Larger leaves would then scale into larger whole plants, which might partly explain the increases in size that accompanied domestication.

Limited evolutionary divergence of seedlings after the domestication of plant species.

The most vulnerable stage in the life of plants is the seedling. The transition from wild to agricultural land that plants experienced during and after domestication implied a noticeable change in the seedlings' environment. Building on current knowledge of seedling ecology, and on previous studies of cassava, we hypothesise that cultivation should have promoted epigeal germination of seedlings, and more exposed and photosynthetic cotyledons. To test this hypothesis, we phenotyped seedling morpho-functional traits in a set of domesticated and wild progenitor accessions of 20 Eudicot herbaceous crop species. Qualitative traits like epi- versus hypogeal germination, leafy versus storage type of cotyledons, or crypto- versus phanerocotyledonar germination, remained conserved during the domestication of all 20 species. Lengths of hypocotyls and epicotyls, of cotyledon petioles, and indices of cotyledon exposure to the aboveground environment changed during evolution under cultivation. However, those changes occurred in diverse directions, depending on the crop species. No common seedling phenotypic convergence in response to domestication was thus detected among the group of species studied here. Also, none of the 20 crops evolved in accordance with our initial hypothesis. Our results reject the idea that strong selective filters exerted unconsciously by artificial selection should have resulted in generalised channelling of seedling morphology towards more productive and more herbivore risky phenotypes. This result opens up unexplored opportunities for directional breeding of seedling traits.

Comparative germination ecology of two altitudinal vicariant Saxifraga species endemic to the north of Spain.

Seeds of high-mountain species are thought to germinate rapidly, synchronously and at high percentages after a cold period, with limited dependence on the external environment; yet, empirical evidence only partially supports this behaviour. We performed a comparative study of the germination response of two closely related taxa along an altitude gradient in northern Spain. Seeds from several maternal families of six populations of Saxifraga trifurcata (lowland species) and S. canaliculata (highland species) were subjected to temperature and stratification treatments. Germination percentages and germination rates were analysed using generalised linear mixed modelling and accelerated failure-time modelling. We found that germination percentages and germination rates were high and dependent on incubation temperature in both species. Within species, seeds from higher altitudes had higher germination percentages under all conditions. Cold-wet stratification negatively affected germination success, particularly in the lowland species. Overall, the highland species was less responsive to the experimental treatments and showed more synchronous germination patterns. We conclude that seeds from these two Saxifraga species germinate as efficiently as species from other habitats, but have a narrower germination response, probably due to the stronger selective pressures in their harsh environments. Finally, a cold, wet stratification period is not a prerequisite for the germination of high-mountain S. canaliculata, and its strong negative effect on the germination of its lowland relative S. trifurcata may contribute to the altitudinal segregation of these two species.

The optimal age for performing surgery on patients with MEN 2B syndrome.

Multiple endocrine neoplasia (MEN) syndromes are characterized by the association of various endocrine neoplasias. Prophylactic thyroidectomy is the treatment of choice for patients with RET gene mutations. The age at which patients undergo prophylactic thyroidectomy may vary depending on the position of the RET gene codon. In cases of MEN 2B, when the mutation is carried in codons 883, 918 or 922, prophylactic thyroidectomy is performed prior to 6 months of age, due to the increased aggressiveness of these heterozygosities, which are capable of determining the onset of medullary cancer during the first months of life. We present two heterozygous twin patients with MEN 2B syndrome who were born 32 weeks premature, and who underwent prophylactic thyroidectomy at 7 months of age. The patients were carriers of the mutation at codon 918. We suggested the early surgery at 7 months as, due to their prematurity, the patients were required to gain weight to improve their condition prior to surgery. The two patients had medullary thyroid carcinoma without lymph node involvement. In conclusion, for a truly prophylactic thyroidectomy, such patients should undergo surgery within the first month of life, particularly if these patients are carriers of the mutation in codons 883, 918 or 922.

Leaf exchange in a Mediterranean shrub: water, nutrient, non-structural carbohydrate and osmolyte dynamics.

Leaf exchange is an abrupt phenological event that drastically modifies the morphology and physiology of the aerial portion of the plant. We examined if water and osmolyte differences between old leaves and new organs trigger leaf exchange, and whether the differences are closely linked to the resource resorption process in senescing leaves. We monitored concentrations of osmolyte, water, non-structural carbohydrate, nitrogen and potassium in senescing leaves and in emerging new leaves and inflorescences of a Mediterranean leaf exchanger (Cistus laurifolius L.) growing in NE Spain. Old leaves rehydrated markedly during most of the senescence process, which co-occurred with the extension of new shoots, suggesting the lack of a clear-cut switch in water supply from old to new organs. The accumulation of osmolytes in the early stage of leaf senescence might account for this rehydration. Osmolyte dynamics in old leaves depended largely on the progression of resource resorption from senescing organs but were mostly unrelated to water content during late senescence. We conclude that dehydration of old leaves is not a prerequisite for the triggering of leaf exchange. The finding that most nutrients and carbohydrates accumulated in new organs before senescing leaves massively exported resources, and the absence of relevant differences between the dynamics of old leaves at the base of inflorescences and those at the base of vegetative shoots, indicate that the nutrient and carbohydrate demands of new organs do not trigger leaf exchange.

Costs of reproduction as related to the timing of phenological phases in the dioecious Shrub Pistacia lentiscus L.

Females of woody dioecious species usually devote more resources to reproduction than males. This may lead to a decrease in female survival and growth. The costs of reproduction, however, can be lightened through a number of mechanisms, as for example avoiding the temporal coincidence of reproduction and vegetative growth. The aim of this study was to evaluate whether males and females of P. lentiscus differ in the timing of their vegetative growth, and to assess whether the sequencing of vegetative growth and reproduction reduces reproductive costs. We monitored phenology in males and females. We also compared male and female allocation of nutrients and biomass in the branch, and the developmental stability of the growing shoots. We did this both prior to and at the end of the fruiting period. Males and females showed similar vegetative and flowering phenologies. Males invested more biomass in flowering, but the sexes showed equal vegetative biomass and nutrient content prior to the fruiting period. In female branches, no trade-off was found between fruit load and current-year vegetative growth. In P. lentiscus, avoiding the overlap of flowering, vegetative growth and fruiting probably contributes to reduce the immediate costs of reproductive efforts, both in males and females.

Relationships between phenology and the remobilization of nitrogen, phosphorus and potassium in branches of eight Mediterranean evergreens.

Few studies have examined the effects of plant growth on nutrient remobilization in phenologically contrasting species. Here we evaluated the consequences of above-ground seasonality of growth and leaf shedding on the remobilization of nutrients from branches in eight evergreen Mediterranean phanaerophytes that differ widely in phenology. Vegetative growth, flower bud formation, flowering, fruiting, leaf shedding, and the variations in nitrogen (N), phosphorus (P) and potassium (K) pools in branches throughout the year were monitored in each species. Nitrogen and P remobilization occurred in summer, after vegetative growth and synchronously with leaf shedding. Despite the time-lag between growth and remobilization, the branches that invested more nutrients in vegetative growth also remobilized more nutrients from their old organs. Potassium remobilization peaked in the climatically harshest periods, and appears to be related to osmotic requirements. We conclude that N and P remobilization occurs mainly associated with leaf senescence, which might be triggered by factors such as the replenishment of nutrient reserves in woody organs, the hormonal relations between new and old leaves, or the constraints that summer drought poses on the amount of leaf area per branch in summer.

Environmental constraints on phenology and internal nutrient cycling in the Mediterranean winter-deciduous shrub Amelanchier ovalis Medicus.

The functional adjustments of winter-deciduous perennials to Mediterranean conditions have received little attention. The objectives of this study were: (i) to determine whether Amelanchier ovalis, a winter-deciduous shrub of Mediterranean and sub-Mediterranean regions, has nutritional and phenological traits in common with temperate zone deciduous trees and shrubs and (ii) to determine the constraints of Mediterranean environmental conditions on these traits. Over two years, phenology and nitrogen, and phosphorus concentrations were monitored monthly in the crown of A. ovalis. Leaf longevity, survival and nutrient resorption from senescing leaves were used to infer nutrient use efficiency and retention times of nutrients within the crown. In A. ovalis, bud burst was much earlier than in temperate deciduous trees and shrubs. Most vegetative and reproductive growth occurred in spring. Limited phenological development took place during the summer drought period. Unexpectedly, leaf shedding was very gradual, which might be related to water shortages in summer. Leaf longevity, nutrient resorption from senescing leaves, and maximum leaf nutrient concentrations indicated that nutrient retention times were short and nutrient use efficiency was low compared to that found in temperate deciduous plants and co-occurring Mediterranean evergreens. A. ovalis exhibited phenological development appropriate for a Mediterranean climate, although its limited ability to retain nutrients likely restricts the types of sites that it can occupy.

Genetic and physical characterization of chromosome 4DL in wheat.

The long arm of chromosome 4D in wheat (Triticum aestivum L.) has been shown in previous studies to harbor genes of agronomic importance. A major dominant gene conferring Aluminum (Al) tolerance (Alt2 in 'Chinese Spring' and AltBH in 'BH 1146'), and the Knal locus controlling the K+/Na+ discrimination in saline environments have been mapped to this chromosome arm. However, accurate information on the genetic and physical location of markers related to any of these genes is not available and would be useful for map-based cloning and marker-assisted plant breeding. In the present study, using a population of 91 recombinant inbred lines segregating for Al tolerance, we provide a more extensive genetic linkage map of the chromosome arm 4DL based on RFLP, SSR, and AFLP markers, delimiting the AltBH gene to a 5.9-cM interval between markers Xgdm125 and Xpsr914. In addition, utilizing a set of wheat deletion lines for chromosome arm 4DL, the AltBH gene was physically mapped to the distal region of the chromosome, between deletion breakpoints 0.70 and 0.86, where the kilobase/centimorgan ratio is assumed to be low, making the map-based cloning of the gene a more realistic goal. The polymorphism rates in chromosome arm 4DL for the different types of markers used were extremely low, as confirmed by the physical mapping of AFLPs. Finally, analysis of 1 Mb of contiguous sequence of Arabidopsis chromosome 5 flanking the gene homologous to the BCD1230 clone (a cosegregating marker in our population coding for a ribulose-5-phosphate-3-epimerase gene), revealed a previously identified region of stress-related and disease-resistance genes. This could explain the collinearity observed in comparative mapping studies among different species and the low level of polymorphism detected in the chromosome arm 4DL in hexaploid wheat.