High viscosity and anisotropy characterize the cytoplasm of fungal dormant stress-resistant spores

Ascospores of the fungus Talaromyces macrosporus are dormant and extremely stress resistant, whereas fungal conidia--the main airborne vehicles of distribution--are not. Here, physical parameters of the cytoplasm of these types of spores were compared. Cytoplasmic viscosity and level of anisotropy as judged by spin probe studies (electron spin resonance) were extremely high in dormant ascospores and during early germination and decreased only partly after trehalose degradation and glucose efflux. Upon prosilition (ejection of the spore), these parameters fell sharply to values characteristic of vegetative cells. These changes occurred without major volume changes that suggest dramatic changes in cytoplasmic organization. Azide reversibly inhibited prosilition as well as the decline in cytoplasmic parameters. No organelle structures were observed in etched, cryoplaned specimens of ascospores by low-temperature scanning electron microscopy (LTSEM), confirming the high cytoplasmic viscosity. However, cell structures became visible upon prosilition, indicating reduced viscosity. The viscosity of fresh conidia of different Penicillium species was lower, namely, 3.5 to 4.8 cP, than that of ascospores, near 15 cP. In addition the level of anisotropic motion was markedly lower in these cells (h(0)/h(+1) = 1.16 versus 1.4). This was confirmed by LTSEM images showing cell structures. The decline of cytoplasmic viscosity in conidia during germination was linked with a gradual increase in cell volume. These data show that mechanisms of cytoplasm conservation during germination differ markedly between ascospores and conidia.     

Natal origin affects host preference and larval performance relationships in a tritrophic system

Many insects face the challenge to select oviposition sites in heterogeneous environments where biotic and abiotic factors can change over time. One way to deal with this complexity is to use sensory experiences made during developmental stages to locate similar habitats or hosts in which larval development can be maximized. While various studies have investigated oviposition preference and larval performance relationships in insects, they have largely overlooked that sensory experiences made during the larval stage can affect such relationships. We addressed this issue by determining the role of natal experience on oviposition preference and larval performance relationships in a tritrophic system consisting of Galerucella sagittariae, feeding on the two host plants Potentilla palustris and Lysimachia thyrsiflora, and its larval parasitoid Asecodes lucens. We firstly determined whether differences in host‐derived olfactory information could lead to divergent host selection, and secondly, whether host preference could result in higher larval performance based on the natal origin of the insects. Our results showed that the natal origin and the quality of the current host are both important aspects in oviposition preference and larval performance relationships. While we found a positive relationship between preference and performance for natal Lysimachia beetles, natal Potentilla larvae showed no such relationship and developed better on L. thyrsiflora. Additionally, the host selection by the parasitoid was mainly affected by the natal origin, while its performance was higher on Lysimachia larvae. With this study, we showed that the relationship between oviposition preference and larval performance depends on the interplay between the natal origin of the female and the quality of the current host. However, without incorporating the full tritrophic context of these interactions, their implication in insect fitness and potential adaptation cannot be fully understood.     

Activation of Raf/ERK1/2 MAP kinase pathway is involved in GM-CSF-induced proliferation and survival but not in erythropoietin-induced differentiation of TF-1 cells

The involvement of MAPK pathways in differentiation, proliferation and survival was investigated by comparing Epo and GM-CSF signalling in human factor-dependent myeloerythroid TF-1 cells with abnormal Epo-R. GM-CSF withdrawal induced cell-cycle arrest and apoptosis accompanied by increased caspase-3 activity, DNA degradation and reduced expression of the antiapoptotic Bcl-2 and Bcl-xl proteins. Readministration of GM-CSF but not Epo reversed these processes and induced proliferation. The GM-CSF promoted cell survival and proliferation correlated with MEK-1 dependent ERK1/2, Elk-1 and CREB phosphorylation and Egr-1, c-Fos expression as well as with increased STAT-5, AP-1, c-Myb and NF-kappaB DNA-binding. In contrast, Epo failed to activate the Raf-1/ERK1/2 MAPK pathway or to induce Egr-1 and/or c-Fos expression, while it induced erythroid differentiation in GM-CSF-deprived cells. In addition, the Epo-induced haemoglobin production was inhibited in the presence of GM-CSF. These results demonstrate that the activation of MAPK cascade is not necessary for Epo-induced haemoglobin production in TF-1 cells and suggest a negative cross-talk between the signalling of GM-CSF-stimulated cell proliferation and Epo-induced erythroid differentiation.     

The polyene antimycotics nystatin and filipin disrupt the plasma membrane, whereas natamycin inhibits endocytosis in germinating conidia of Penicillium discolor

Aims:  To investigate the differences in membrane permeability and the effect on endocytosis of the polyene antimycotics nystatin, filipin and natamycin on germinating fungal conidia.
Methods and Results:  The model system was Penicillium discolor , a food spoilage fungus. Filipin resulted in permeabilization of germinating conidia for the fluorescent probes TOTO-1 and FM4-64, but not for ferricyanide ions. Nystatin caused influx of all these compounds while natamycin did not. Untreated germinating conidia internalize the endocytic marker FM4-64. Pretreatment of germinating conidia with natamycin showed a dose and time dependent inhibition of endocytosis as judged by the lack of formation of early endosomal compartments.
Conclusions:  The results obtained from this study indicated that, unlike nystatin and filipin, natamycin is unable to permeabilize germinating conidia, but interferes with endocytosis in a dose and time dependent manner.
Significance and Impact of the Study:  Natamycin acts via a different mode of action than other polyene antimycotics. These results offer useful information for new strategies to prevent fungal spoilage on food products and infection on agricultural crops. For laboratory use, natamycin can be used as a specific inhibitor of early endocytosis in fungal cells.     

Association between PrP genotypes and performance traits in a Welsh Mountain flock

The UK national scrapie plan (NSP) for sheep is based on selection for the resistant ARR/ARR genotype and elimination of susceptible types of the ovine prion protein (PrP) gene. The aim of this study was to estimate the possible association of the PrP genotype and performance traits by using data from the CAMDA Welsh Mountain flock. Four alleles (ARH, ARQ, ARR and VRQ) and 10 genotypes covering all five NSP risk groups were present in the CAMDA flock. Overall, the most common allele was ARR (35.2%), and VRQ was the least common (5.4%). The commonest genotypes were ARR/ARQ (23.7%) and ARR/AHQ (23.1%). The most resistant genotype, ARR/ARR, and the most susceptible genotype, VRQ/VRQ, were found in 10.2% and 0.3%, respectively, of the population tested. The associations of PrP genotypes with weight and ultrasonically scanned traits were investigated in three analyses, the first using genotypes, the second using risk categories and the third using number of alleles. These associations were evaluated by univariate analysis of each trait using an animal model with maternal effects where appropriate, and PrP was included as a fixed effect. Selection for scrapie resistance will not adversely affect progress in the traits considered and is consistent with improvements in muscle depth.     

Artichoke Leaf Morphology and Surface Features in Different Micropropagation Stages

Artichoke (Cynara scolymus L.) leaf size and shape, glandular and covering trichomes, stomatal density, stomata shape, pore area and epicuticular waxes during micropropagation stages were studied by scanning electron microscopy (SEM) and morphometric analysis with the aim to improve the survival rate after transfer to greenhouse conditions. Leaves from in vitro shoots at the proliferation stage showed a spatular shape, ring-shaped stomata, a large number of glandular trichomes and juvenile covering hairs, but failed to show any epicuticular waxes. Leaves from in vitro plants at the root elongation stage showed a lanceolated elliptic shape with a serrated border, elliptical stomata, decreased pore area percentage, stomatal density, and mature covering trichomes. One week after transfer to ex vitro conditions, epicuticular waxes appeared on the leaf surface and stomata and pore area were smaller as compared to in vitro plants. Artichoke acclimatization may be improved by hormonal stimulation of root development, since useful morphological changes on leaves occurred during root elongation.     

Structural differences between hyperhydric and normal in vitro shoots of <Emphasis Type=Italic>Handroanthus impetiginosus</Emphasis> (Mart. ex DC) Mattos (Bignoniaceae)

The anatomy of normal and hyperhydric shoots (leaves and stems) of in vitro Handroanthus impetiginosus was compared using light microscopy, scanning electron microscopy, and transmission electron microscopy. In contrast to normal shoots, hyperhydric shoots presented numerous anatomical abnormalities at the proliferation stage. Disorganized cortex, epidermal holes, epidermal discontinuity, collapsed cells, and other structural characteristics were observed in hyperhydric shoots. So, by using anatomical analysis of in vitro H. impetiginosus shoots at the proliferation stage, we can predict which plants will survive the rhizogenesis and acclimatization stages.     

The most heat-resistant conidia observed to date are formed by distinct strains of Paecilomyces variotii

Fungi colonize habitats by means of spores. These cells are stress-resistant compared with growing fungal cells. Fungal conidia, asexual spores, formed by cosmopolitan fungal genera like Penicillium, Aspergillus and Peacilomyces are dispersed by air. They are present in places where food products are stored and as a result, they cause food spoilage. Here, we determined the heterogeneity of heat resistance of conidia between and within strains of Paecilomyces variotii, a spoiler of foods such as margarine, fruit juices, canned fruits and non-carbonized sodas. Out of 108 strains, 31 isolates showed a conidial survival >10% after a 10-min-heat treatment at 59°C. Three strains with different conidial heat resistance were selected for further phenotyping. Conidia of DTO 212-C5 and DTO 032-I3 showed 0.3% and 2.6% survival in the screening respectively, while survival of DTO 217-A2 conidia was >10%. The decimal reduction times of these strains at 60°C (D₆₀ value) were 3.7 ± 0.08, 5.5 ± 0.35 and 22.9 ± 2.00 min respectively. Further in-depth analysis revealed that the three strains showed differences in morphology, spore size distributions, compatible solute compositions and growth under salt stress. Conidia of DTO 217-A2 are the most heat-resistant reported so far. The ecological consequences of this heterogeneity of resistance, including food spoilage, are discussed.