Should food‐deceptive species flower before or after rewarding species? An experimental test of pollinator visitation behaviour under contrasting phenologies

Many plant species reward their pollinators, whereas some species, particularly among orchids, do not. Similarity of floral cues between co‐flowering species influences how rapidly pollinators learn to avoid deceptive plants. This learning process, which affects the reproductive success of deceptive plants, may additionally depend on relative timing of flowering of sympatric rewarding and deceptive species. We tested the combined effects of corolla colour similarity and flowering order of rewarding and deceptive artificial inflorescences on visitation by naïve bumblebees. When deceptive inflorescences were offered after rewarding inflorescences, bumblebees visited them four times more often if both species were similar compared with when they were dissimilar. Pollinator visitation rate to deceptive inflorescences offered before rewarding inflorescences was intermediate and independent of similarity. Thus, early‐flowering deceptive species avoid the costs of dissimilarity with rewarding species. This mechanism may favour adaptive evolution of flowering phenology in deceptive species and explain why temperate deceptive orchids usually flower earlier than rewarding ones.     

Serological detection of Plasmodium vivax malaria using recombinant proteins corresponding to the 19-kDa C-terminal region of the merozoite surface protein-1

BACKGROUND: Serological tests to detect antibodies specific to Plasmodium vivax could be a valuable tool for epidemiological studies, for screening blood donors in areas where the malaria is not endemic and for diagnosis of infected individuals. Because P. vivax cannot be easily obtained in vitro, ELISA assays using total or semi-purified antigens are rarely used. Based on this limitation, we tested whether recombinant proteins representing the 19 kDa C-terminal region of the merozoite surface protein-1 of P. vivax (MSP119) could be useful for serological detection of malaria infection. METHODS: Three purified recombinant proteins produced in Escherichia coli (GST-MSP119, His6-MSP119 and His6-MSP119-PADRE) and one in Pichia pastoris (yMSP119-PADRE) were compared for their ability to bind to IgG antibodies of individuals with patent P. vivax infection. The method was tested with 200 serum samples collected from individuals living in the north of Brazil in areas endemic for malaria, 53 serum samples from individuals exposed to Plasmodium falciparum infection and 177 serum samples from individuals never exposed to malaria. RESULTS: Overall, the sensitivity of the ELISA assessed with sera from naturally infected individuals was 95%. The proportion of serum samples that reacted with recombinant proteins GST-MSP119, His6-MSP119, His6-MSP119-PADRE and yMSP119-PADRE was 90%, 93.5%, 93.5% and 93.5%, respectively. The specificity values of the ELISA determined with sera from healthy individuals and from individuals with other infectious diseases were 98.3% (GST-MSP119), 97.7% (His6-MSP119 and His6-MSP119-PADRE) or 100% (yMSP119-PADRE). CONCLUSIONS: Our study demonstrated that for the Brazilian population, an ELISA using a recombinant protein of the MSP119 can be used as the basis for the development of a valuable serological assay for the detection of P. vivax malaria.     

The potential for floral mimicry in rewardless orchids: an experimental study

More than one-third of orchid species do not provide their pollinators with either pollen or nectar rewards. Floral mimicry could explain the maintenance of these rewardless orchid species, but most rewardless orchids do not appear to have a rewarding plant that they mimic specifically. We tested the hypothesis that floral mimicry can occur through similarity based on corolla colour alone, using naive bumble-bees foraging on arrays of plants with one rewarding model species, and one rewardless putative mimic species (Dactylorhiza sambucina) which had two colour morphs. We found that when bees were inexperienced, they visited both rewardless morphs randomly. However, after bees had gained experience with the rewarding model, and it was removed from the experiment, bees resampled preferentially the rewardless morph most similar to it in corolla colour. This is the first clear evidence, to our knowledge, that pollinators could select for floral mimicry. We suggest that floral mimicry can be a selective force acting on rewardless orchids, but only under some ecological conditions. In particular, we argue that selection on early-flowering rewardless orchids that receive visits from a large pool of naive pollinators will be weakly influenced by mimicry.     

Morphogenic Regulation of Pathotoxin Synthesis in Cochliobolus carbonum

The fungus Cochliobolus carbonum causes leaf spot disease of maize. Highly virulent isolates of the pathogen produce a host-selective, peptide toxin that is active against susceptible genotypes of maize. Prior to infection, spores must germinate and differentiate appressoria, structures specialized for leaf penetration. Analysis of spore germination fluids by plasma desorption mass spectrometry, which allowed detection of as little as 0.5 ng toxin, revealed that spores induced to form appressoria in vitro synthesized and released the toxin at a time coincident with maturation of appressoria. Spores incubated under conditions that did not induce appressorium formation failed to produce toxin. These observations indicate that synthesis of the host-selective toxin, which is essential for successful pathogenesis of maize by C. carbonum, is regulated by infection-related morphogenesis.     

Inhibition of matrix metalloproteinase-9 secretion by dimethyl sulfoxide and cyclic adenosine monophosphate in human monocytes

BACKGROUND Matrix metalloproteinases(MMPs),including MMP-9,are an integral part of the immune response and are upregulated in response to a variety of stimuli.New details continue to emerge concerning the mechanistic and regulatory pathways that mediate MMP-9 secretion.There is significant evidence for regulation of inflammation by dimethyl sulfoxide(DMSO)and 3',5'-cyclic adenosine monophosphate(cAMP),thus investigation of how these two molecules may regulate both MMP-9 and tumor necrosis factorα(TNFα)secretion by human monocytes was of high interest.The hypothesis tested in this study was that DMSO and cAMP regulate MMP-9 and TNFαsecretion by distinct mechanisms.AIM To investigate the regulation of lipopolysaccharide(LPS)-stimulated MMP-9 and tumor necrosis factorαsecretion in THP-1 human monocytes by dimethyl sulfoxide and cAMP.METHODS The paper describes a basic research study using THP-1 human monocyte cells.All experiments were conducted at the University of Missouri-St.Louis in the Department of Chemistry and Biochemistry.Human monocyte cells were grown,cultured,and prepared for experiments in the University of Missouri-St.Louis Cell Culture Facility as per accepted guidelines.Cells were treated with LPS for selected exposure times and the conditioned medium was collected for analysis of MMP-9 and TNFαproduction.Inhibitors including DMSO,cAMP regulators,and anti-TNFαantibody were added to the cells prior to LPS treatment.MMP-9 secretion was analyzed by gel electrophoresis/western blot and quantitated by ImageJ software.TNFαsecretion was analyzed by enzyme-linked immuno sorbent assay.All data is presented as the average and standard error for at least 3 trials.Statistical analysis was done using a two-tailed paired Student t-test.P values less than 0.05 were considered significant and designated as such in the Figures.LPS and cAMP regulators were from Sigma-Aldrich,MMP-9 standard and antibody and TNFαantibodies were from R&D Systems,and amyloid-βpeptide was from rPeptide.RESULTS In our investigation of MMP-9 secretion from THP-1 human monocytes,we made the following findings.Inclusion of DMSO in the cell treatment inhibited LPSinduced MMP-9,but not TNFα,secretion.Inclusion of DMSO in the cell treatment at different concentrations inhibited LPS-induced MMP-9 secretion in a dosedependent fashion.A cell-permeable cAMP analog,dibutyryl cAMP,inhibited both LPS-induced MMP-9 and TNFαsecretion.Pretreatment of the cells with the adenylyl cyclase activator forskolin inhibited LPS-induced MMP-9 and TNFαsecretion.Pretreatment of the cells with the general cAMP phosphodiesterase inhibitor IBMX reduced LPS-induced MMP-9 and TNFαin a dose-dependent fashion.Pre-treatment of monocytes with an anti-TNFαantibody blocked LPSinduced MMP-9 and TNFαsecretion.Amyloid-βpeptide induced MMP-9 secretion,which occurred much later than TNFαsecretion.The latter two findings strongly suggested an upstream role for TNFαin mediating LPS-stimulate MMP-9 secretion.CONCLUSION The cumulative data indicated that MMP-9 secretion was a distinct process from TNFαsecretion and occurred downstream.First,DMSO inhibited MMP-9,but not TNFα,suggesting that the MMP-9 secretion process was selectively altered.Second,cAMP inhibited both MMP-9 and TNFαwith a similar potency,but at different monocyte cell exposure time points.The pattern of cAMP inhibition for these two molecules suggested that MMP-9 secretion lies downstream of TNFαand that TNFαmay a key component of the pathway leading to MMP-9 secretion.This temporal relationship fit a model whereby early TNFαsecretion directly led to later MMP-9 secretion.Lastly,antibody-blocking of TNFαdiminished MMP-9 secretion,suggesting a direct link between TNFαsecretion and MMP-9 secretion.