Relatedness among honeybees (Apis mellifera) of a drone congregation

The honeybee (Apis mellifera) queen mates during nuptial flights, in the so-called drone congregation area where many males from surrounding colonies gather. Using 20 highly polymorphic microsatellite loci, we studied a sample of 142 drones captured in a congregation close to Oberursel (Germany). A parentage test based on lod score showed that this sample contained one group of four brothers, six groups of three brothers, 20 groups of two brothers and 80 singletons. These values are very close to a Poisson distribution. Therefore, colonies were apparently equally represented in the drone congregation, and calculations showed that the congregation comprised males that originated from about 240 different colonies. This figure is surprisingly high. Considering the density of colonies around the congregation area and the average flight range of males, it suggests that most colonies within the recruitment perimeter delegated drones to the congregation with an equal probability, resulting in an almost perfect panmixis. Consequently, the relatedness between a queen and her mates, and hence the inbreeding coefficient of the progeny, should be minimized. The relatedness among the drones mated to the same queen is also very low, maximizing the genetic diversity among the different patrilines of a colony.     

Ovipositing female house flies provision offspring larvae with bacterial food

Symbiotic bacteria on house fly eggs, Musca domestica L. (Diptera: Muscidae), provide ovipositional cues for conspecific female flies and curtail the growth of fungi that compete with fly larval offspring for resources. Because bacteria are also essential dietary constituents for developing larvae, we tested the hypothesis that egg‐derived bacteria support development of larvae to adults. From house fly eggs, we isolated and identified 12 strains of bacteria, eight and four of which were previously shown to induce and inhibit oviposition, respectively. When larvae were provisioned with a total dose of 10⁶–10⁷ colony‐forming units of bacteria from either the oviposition‐inducing or inhibiting group, or from both groups together, significantly more larvae completed development. Thus, egg‐associated bacteria could be a fail‐safe mechanism that ensures a bacterial food supply for larval offspring, particularly if the resource selected by parent females is poor in bacterial food.     

The pathogen causing Dutch elm disease makes host trees attract insect vectors

Dutch elm disease is caused by the fungal pathogen Ophiostoma novo-ulmi which is transmitted by the native elm bark beetle, Hylurgopinus rufipes. We have found that four semiochemicals (the monoterpene (-)-beta-pinene and the sesquiterpenes (-)-alpha-cubebene, (+)-spiroaxa-5,7-diene and (+)-delta-cadinene) from diseased American elms, Ulmus americana, synergistically attract H. rufipes, and that sesquiterpene emission is upregulated in elm trees inoculated with O. novo-ulmi. The fungus thus manipulates host trees to enhance their apparency to foraging beetles, a strategy that increases the probability of transportation of the pathogen to new hosts.     

High-throughput SNP genotyping by single-tube PCR with Tm-shift primers

Despite many recent advances in high-throughput single nucleotide polymorphism (SNP) genotyping technologies, there is still a great need for inexpensive and flexible methods with a reasonable throughput. Here we report substantial modifications and improvements to an existing homogenous allele-specific PCR-based SNP genotyping method, making it an attractive new option for researchers engaging in candidate gene studies or following up on genome-wide scans. In this advanced version of the melting temperature (Tm)-shift SNP genotyping method, we attach two GC-rich tails of different lengths to allele-specific PCR primers, such that SNP alleles in genomic DNA samples can be discriminated by the Tms of the PCR products. We have validated 306 SNP assays using this method and achieved a success rate in assay development of greater than 83% under uniform PCR conditions. We have developed a standalone software application to automatically assign genotypes directly from melting curve data. To demonstrate the accuracy of this method, we typed 592 individuals for 6 SNPs and showed a high call rate (>98%) and high accuracy (>99.9%). With this method, 6-10,000 samples can be genotyped per day using a single 384-well real-time thermal cycler with 2-4 standard 384-well PCR instruments.     

Anxiolytic-like action in mice treated with nitrous oxide and oral triazolam or diazepam

Few animal studies have explored the interaction of nitrous oxide (N2O) with a benzodiazepine (BNZ) administered by the oral route, as used in clinical procedures involving "conscious sedation". The purpose of this study was to evaluate the relative "anxiolytic-like" and sedative effectiveness of N2O, oral triazolam (TRIAZ; Halcion) or oral diazepam (DIAZ; Valium), either alone or in various combinations of drugs and doses. One hundred and twelve Swiss Webster male mice, 35-45 days old, were assigned to 28 groups, each of which contained four mice. The mouse staircase test was used for the assessment of anxiety (number of rearings) and sedation (number of steps ascended). Three doses of oral TRIAZ (0.1, 0.3, 1.0 mg/kg) or DIAZ (2.0, 3.5, 5.0 mg/kg) were given in combination with room air, or N2O/O2 at a N2O concentration of 25, 50 or 75%. Each mouse was tested once. N2O alone did not reduce NR in any concentration, but caused a significant increase in locomotion. DIAZ without N2O reduced NR only with the middle and high doses, but the addition of N2O significantly enhanced the anxiolytic-like effect of all DIAZ doses. TRIAZ, alone, reduced NR only in the highest dose, but added N2O resulted in anxiolytic-like behavior with all three TRIAZ doses. The sedative effects of the BNZs were extremely variable. Only the middle dose of DIAZ plus 25% N2O unequivocally reduced the number of steps ascended, i.e., caused sedation. TRIAZ lacked the inverted U-shaped dose-response relationship with NR usually seen with DIAZ. TRIAZ, therefore, provides better dose control. This behavioral animal model indicates that the optimal combinations for reduction of anxiety-like behavior with minimal effects on sedation are 0.1 mg/kg oral TRIAZ with 25% N2O or 2.0 mg/kg oral DIAZ with 25% N2O.     

Leucine-specific, functional interactions between human immunodeficiency virus type 1 Nef and adaptor protein complexes

The human immunodeficiency virus type 1 virulence protein Nef interacts with the endosomal sorting machinery via a leucine-based motif. Similar sequences within the cytoplasmic domains of cellular transmembrane proteins bind to the adaptor protein (AP) complexes of coated vesicles to modulate protein traffic, but the molecular basis of the interactions between these motifs and the heterotetrameric complexes is controversial. To identify the target of the Nef leucine motif, the native sequence was replaced with either leucine- or tyrosine-based AP-binding sequences from cellular proteins, and the interactions with AP subunits were correlated with function. Tyrosine motifs predictably modulated the interactions between Nef and the mu subunits of AP-1, AP-2, and AP-3; heterologous leucine motifs caused little change in these interactions. Conversely, leucine motifs mediated a ternary interaction between Nef and hemicomplexes containing the sigma1 plus gamma subunits of AP-1 or the sigma3 plus delta subunits of AP-3, whereas tyrosine motifs did not. Similarly, only leucine motifs supported the Nef-mediated association of AP-1 and AP-3 with endosomal membranes in cells treated with brefeldin A. Functionally, Nef proteins containing leucine motifs down-regulated CD4 from the cell surface and enhanced viral replication, whereas those containing tyrosine motifs were inactive. Apparently, the interaction of Nef with the mu subunits of AP complexes is insufficient for function. A leucine-specific mode of interaction that likely involves AP hemicomplexes is further required for Nef activity. The mu and hemicomplex interactions may cooperate to yield high avidity binding of AP complexes to Nef. This binding likely underlies the unusual ability of Nef to induce the stabilization of these complexes on endosomal membranes, an activity that correlates with enhancement of viral replication.     

Sexual reproduction in the tropical corallimorpharian Rhodactis rhodostoma

Abstract. Polyps of the tropical corallimorpharian Rhodactis rhodostoma segregate sexes between center and edge positions within aggregations produced by clonal replication. On a reef flat at Eilat, northern Red Sea, infertile polyps and males occur mainly along the edges of clonal aggregations, while females mostly occupy central positions within each aggregation. In addition, on the inner to middle reef flat where polyps of this species are abundant, aggregations consist mostly of females. On the outer reef flat, where polyps are rare, a sampled aggregation consisted mostly of males and infertile polyps. Male polyps are significantly smaller than females, and the smallest polyps are infertile. Fecundity increases significantly with polyp size in females, but testis size and number do not vary with body size in males. Oocytes are present in polyps during most of the year and gradually increase in size until annual spawning in June-July during the period of maximum day length. Testes do not vary significantly in size during the year and remain a small proportion of body mass (>8%). In contrast, females invest up to 30% of their body mass into gonads during the months immediately before spawning. The annual spawning of gametes coincides with a temporary drop in the frequency of clonal replication by polyps. We estimate that each female polyp of R. rhodostoma may release up to 3000 large eggs (500 μm in maximum diameter) each summer. The high investment of this corallimorpharian in sexual production of planktonic propagules may allow rapid dispersal to reef habitats distant from parent populations.     

Maintenance of the enteric stem cell niche by bacterial lipopolysaccharides? Evidence and perspectives

The enteric nervous system (ENS) has to respond to continuously changing microenvironmental challenges within the gut and is therefore dependent on a neural stem cell niche to keep the ENS functional throughout life. In this study, we hypothesize that this stem cell niche is also affected during inflammation and therefore investigated lipopolysaccharides (LPS) effects on enteric neural stem/progenitor cells (NSPCs). NSPCs were derived from the ENS and cultured under the influence of different LPS concentrations. LPS effects upon proliferation and differentiation of enteric NSPC cultures were assessed using immunochemistry, flow cytometry, western blot, Multiplex ELISA and real‐time PCR. LPS enhances the proliferation of enteric NSPCs in a dose‐dependent manner. It delays and modifies the differentiation of these cells. The expression of the LPS receptor toll‐like receptor 4 on NSPCs could be demonstrated. Moreover, LPS induces the secretion of several cytokines. Flow cytometry data gives evidence for individual subgroups within the NSPC population. ENS‐derived NSPCs respond to LPS in maintaining at least partially their stem cell character. In the case of inflammatory disease or trauma where the liberation and exposure to LPS will be increased, the expansion of NSPCs could be a first step towards regeneration of the ENS. The reduced and altered differentiation, as well as the induction of cytokine signalling, demonstrates that the stem cell niche may take part in the LPS‐transmitted inflammatory processes in a direct and defined way.