Medicago truncatula IPD3 is a member of the common symbiotic signaling pathway required for rhizobial and mycorrhizal symbioses

Legumes form endosymbiotic associations with nitrogen-fixing bacteria and arbuscular mycorrhizal (AM) fungi which facilitate nutrient uptake. Both symbiotic interactions require a molecular signal exchange between the plant and the symbiont, and this involves a conserved symbiosis (Sym) signaling pathway. In order to identify plant genes required for intracellular accommodation of nitrogen-fixing bacteria and AM fungi, we characterized Medicago truncatula symbiotic mutants defective for rhizobial infection of nodule cells and colonization of root cells by AM hyphae. Here, we describe mutants impaired in the interacting protein of DMI3 (IPD3) gene, which has been identified earlier as an interacting partner of the calcium/calmodulin-dependent protein, a member of the Sym pathway. The ipd3 mutants are impaired in both rhizobial and mycorrhizal colonization and we show that IPD3 is necessary for appropriate Nod-factor-induced gene expression. This indicates that IPD3 is a member of the common Sym pathway. We observed differences in the severity of ipd3 mutants that appear to be the result of the genetic background. This supports the hypothesis that IPD3 function is partially redundant and, thus, additional genetic components must exist that have analogous functions to IPD3. This explains why mutations in an essential component of the Sym pathway have defects at late stages of the symbiotic interactions.     

Individual vocal production in a sperm whale (Physeter macrocephalus) social unit

The vocal repertoires of group‐living animals may communicate individual or group identity. Female and juvenile sperm whales live in long‐term social units that can be assigned to vocal clans based on the pattern of clicks in coda vocalizations. An unusual set of circumstances allowed us to record the vocalizations of photo‐identified individuals within a single social unit over a 41 d period. Using click interpulse intervals, we were able to assign codas to individuals and investigate coda production at the individual level within a social unit for the first time. Adult females in the unit vocalized at approximately equal rates. A calf and juvenile, both male, vocalized less often than the adult females. Repertoires were indistinguishable for all unit members apart from a mother and her calf, which possessed significantly different repertoires—even from one another. We suggest that similarity among the coda repertoires of most unit members indicates a function in advertising unit identity. In contrast, the distinctive repertoires of the calf and its mother may facilitate reunions between these whales. We hypothesize that sperm whales may be able to vary their vocal repertoires as their reproductive status alters the trade‐off between the benefits of individual and group identification.     

S-genotyping of old apple cultivars from the Carpathian basin: methodological, breeding and evolutionary aspects

Apple exhibits self-incompatibility controlled by the multiallelic S-locus. Twenty-three old apple cultivars were S-genotyped using three different approaches (allele-specific polymerase chain reaction (PCR) + cleaved amplified polymorphic sequences (CAPS), consensus PCR + sequencing and consensus PCR + CAPS) to compare the robustness and reliability of these techniques and characterise genotypes from the Carpathian basin that might be useful in resistance breeding. Best results were obtained using the ASPF3 and ASPR3S consensus primer pair that detected 96% of all alleles carried by the 23 cultivars tested. Flow cytometry analysis was also needed to control the completeness of the genotypes as was seen in case of a tetraploid cultivar with only three assigned S-alleles. The genetic disparity between the old Carpathian basin and modern apple cultivars was indicated by differences in allele frequency data (S ₉, S ₂₄ and S ₂₆) as well as single nucleotide polymorphisms in S ₁, S ₂, S ₇ S ₂₄ and S ₂₆ and indels in S ₂₀ and S ₂₆ alleles. An alignment of partial genomic sequences indicated trans-specific and trans-generic evolution of S-ribonuclease alleles in the Maloideae subfamily (S ₂₆ and S ₂₈) and a possibly recent introgression event (S ₁) between Malus × domestica and Malus sylvestris. These data suggest that the genome of old cultivars from the Carpathian basin was enriched by several Malus taxa and are free from the consequences of modern breeding. These cultivars may contribute to the widening of the genetic basis of cultivated apple and prevent genetic erosion in future commercial cultivars.     

Superinfection in malaria: Plasmodium shows its iron will

After the bite of a malaria-infected mosquito, the Plasmodium sporozoite infects liver cells and produces thousands of merozoites, which then infect red blood cells, causing malaria. In malaria-endemic areas, several hundred infected mosquitoes can bite an individual each year, increasing the risk of superinfection. However, in infants that are yet to acquire immunity, superinfections are infrequent. We have recently shown that blood-stage parasitaemia, above a minimum threshold, impairs the growth of a subsequent sporozoite infection of liver cells. Blood-stage parasites stimulate the production of the host iron-regulatory factor hepcidin, which redistributes iron away from hepatocytes, reducing the development of the iron-dependent liver stage. This could explain why Plasmodium superinfection is not often found in young nonimmune children. Here, we discuss the impact that such protection from superinfection might have in epidemiological settings or in programmes for controlling malaria, as well as how the induction of hepcidin and redistribution of iron might influence anaemia and the outcome of non-Plasmodium co-infections.     

Identifying gene targets for the metabolic engineering of lycopene biosynthesis in Escherichia coli

The identification of genetic targets that are effective in bringing about a desired phenotype change is still an open problem. While random gene knockouts have yielded improved strains in certain cases, it is also important to seek the guidance of cell-wide stoichiometric constraints in identifying promising gene knockout targets. To investigate these issues, we undertook a genome-wide stoichiometric flux balance analysis as an aid in discovering putative genes impacting network properties and cellular phenotype. Specifically, we calculated metabolic fluxes such as to optimize growth and then scanned the genome for single and multiple gene knockouts that yield improved product yield while maintaining acceptable overall growth rate. For the particular case of lycopene biosynthesis in Escherichia coli, we identified such targets that we subsequently tested experimentally by constructing the corresponding single, double and triple gene knockouts. While such strains are suggested (by the stoichiometric calculations) to increase precursor availability, this beneficial effect may be further impacted by kinetic and regulatory effects not captured by the stoichiometric model. For the case of lycopene biosynthesis, the so identified knockout targets yielded a triple knockout construct that exhibited a nearly 40% increase over an engineered, high producing parental strain.     

Formation of trans-verbenol and verbenone from alpha-pinene catalysed by immobilised Picea abies cells

Both enantiomers and the racemate of alpha-pinene were transformed by Picea abies cells immobilised on alginate. The main products were cis- and trans-verbenol, the later being further transformed to verbenone. The enantiomeric purity of each product more or less corresponded to that of the substrate. Transformation by free cells was faster than that by the immobilised cells. The ratio of products differed to some extent between the transformation by free and immobilised cells.     

Regeneration Capacity of Calli Derived from Immature Embryos in Spring Barley Cultivars

Callogenesis, somatic embryogenesis and regeneration capacity in twenty-three agronomically important spring barley (Hordeum vulgare L.) cultivars on induction media with 2,4-dichlorophenoxyacetic acid (2,4-D) or 3,6-dichloro-o-anisic acid (dicamba) and on modified regeneration media were studied. The frequency of zygotic embryos exhibiting callogenesis varied from 88 to 100 % according to genotype. Dicamba was more suitable for somatic embryogenesis induction and exhibited a higher frequency of regenerants than did 2,4-D. Green regenerants were obtained in all cultivars, and there were no albino plants. Except for cv. Victor all cultivars used in the experiment showed lower regeneration capacity as compared to the model cv. Golden Promise. This revised version was published online in July 2006 with corrections to the Cover Date.     

Effect of candidate vaginally-applied microbicide compounds on recognition of antigen by CD4+ and CD8+ T lymphocytes

Vaginally applied antimicrobial compounds (microbicides) are being developed as an alternative method for preventing the spread of sexually transmitted diseases. In addition to identifying compounds effective against a spectrum of sexually transmitted pathogens, it will be important to ensure that these compounds are safe. Avoiding toxicity, inflammatory responses, or alteration of the function of resident immune cells are important considerations for the development of vaginally applied microbicides. Studies were performed with two classes of candidate microbicide compounds to determine if they would interfere with the recognition of antigen by CD4(+) and CD8(+) T lymphocytes. The presence of nontoxic concentrations of the anionic detergent cholic acid or the sulfated polymer lambda carrageenan did not inhibit recognition of immune peptide by antigen-specific T cells. However, antigen recognition by both CD4(+) and CD8(+) T lymphocytes was inhibited in the presence of the naphthalene sulfonate polymer PRO 2000. Brief (4-h) exposure of antigen-presenting cells or T cells to PRO 2000 did not result in inhibition of antigen uptake and processing by antigen-presenting cells or the ability of specific T cells to respond to antigen stimulation, suggesting that the inhibition was temporary. Binding of antibodies specific for CD18, CD8, and CD3 was impaired in the presence of PRO 2000, suggesting that the mechanism by which this microbicide inhibits T cell recognition of antigenic peptide may involve masking or internalization of surface proteins involved in T cell signaling or stabilizing T cell-antigen-presenting cell interactions. The assays described in this study represent a useful means to screen candidate topical microbicide compounds for inappropriate interactions with immune cells and may be useful for prioritization of candidate microbicide compounds.