Magnetic Separation of Phagosomes of Defined Age from Tetrahymena thermophila

ABSTRACT. We describe a new mass isolation procedure for both pure and stage-specific phagosomes from Tetrahymena thermophila . We prepared magnetic iron dextran particles about 1 μm in diameter to label the phagosomes. The oral apparatus of the cells concentrated these particles so readily that after 1 min the majority of the cells had formed a single phagosome. A short wash removed non-ingested particles, enabling us to follow the age-dependent changes of a single labeled phagosome through the cell. Phagosomes of different ages, including very young and nascent phagosomes, were removed easily from the non-magnetic cell debris of mechanically homogenized cells by means of a permanent magnet. The isolated phagosomes are pure as tested by enzymatic assays and light and electron microscopy. Since the yield of pure phagosomes of all ages is high (∼ 90%), this method could be generally applied for phagosome isolation from ciliates.     

Does the decline of gastropods in deep water herald ecosystem change in Lakes Malawi and Tanganyika?

1. Ancient, deep lakes have traditionally been considered as stable, ecological islands, well buffered from environmental change because of their great depth. However, they are not immune to anthropogenic and climatic stress. Ecosystems of the permanently stratified warm Lakes Malawi and Tanganyika in the Great East African Rift are particularly delicate. Their stratification regime has historically limited the distribution of benthic biota to a ‘bathtub ring of biodiversity’, namely substrata in the upper, oxygenated water layer. 2. We use historical data on the endemic deep‐water molluscs of these lakes to assess present‐day stress on their benthic ecosystems. During the 20th century, these molluscs have probably decreased in abundance and migrated to shallower water. 3. These apparent trends have a significance beyond species‐based conservation, foremost because deep‐water organisms heavily rely on the position and temporal stability of the oxycline and therefore provide an early warning of large‐scale changes in the distribution of dissolved oxygen. Oxygen demands have increased in the East African Great Lakes over the last century whereas ventilation of deep water has remained the same or declined. 4. The combination of these factors is resulting in a narrowing of the ring of biodiversity and a changed nutrient flux through this ring. Reduction in the habitat available inevitably puts biota at risk, whereas changes in nutrient flux may cause shifts in the entire ecosystem or the collapse of parts of it. 5. Considering the socioeconomic value of these lakes and the potentially grave implications for their faunal biodiversity and entire ecosystems, existing evidence of faunal decline, especially in taxa that depend strongly on the stratification regime, is of great concern. Moreover, because the factors responsible are widespread and include surface‐water warming, increased run‐off and eutrophication, respiration stress may also develop in other tropical and subtropical lakes.     

Immunocompetent Infants as a Human Reservoir for Pneumocystis jirovecii: Rapid Screening by Non-Invasive Sampling and Real-Time PCR at the Mitochondria1 Large Subunit rRNA Gene

ABSTRACT. We tested a real-time PCR assay targeting the Pneumocystis jirovecii mitochondria1 large subunit rRNA gene on 240 archival nasopharyngeal aspirates from non-immunosuppressed infants. The sensitivity of this assay appeared close to that of a conventional nested-PCR assay targeting the same locus. Because of its one-step procedure, and its sensitivity and rapidity, the real-time PCR assay is particularly suitable for screening individuals parasitized by P. jirovecii within large populations.     

The retrolateral tibial apophysis in spiders—shaped by sexual selection?

The functional significance of the retrolateral tibial apophysis (rta) on the male pedipalps in four spider species with different mating positions is investigated with the help of histological serial sections prepared after freeze-fixing copulating pairs with liquid nitrogen. The results of this study, as well as most data in previous works, suggest that the rta is mostly used to fix the male pedipalp to the female epigyne in order to ensure the intromission of the sperm transferring embolus. This is in accordance with the female choice hypothesis on genitalia which predicts that species-specific genital structures should directly or indirectly contact the female during copulation and thus be shaped by sexual selection.     

Neurophysiological and Behavioral Changes in Non-Target Wildlife Exposed to Organophosphate and Carbamate Pesticides: Thermoregulation, Food Consumption, and Reproduction

Originally utilized or developed as human poisons, anticholinesterasecompounds are among the most widely used pesticides in the worldand non-target wildlife are frequently exposed. Because thesecompounds primarily act by inhibiting acetylcholinesterase atsynapses within the central and peripheral nervous systems,the potential for altering physiological and behavioral responsesessential for survival and reproduction in exposed animals isgreat. We review the effects of acute but sublethal exposureto organophosphates and carbamates on thermoregulation (hypothermia),food consumption (anorexia and altered foraging behavior), andreproduction (altered hormone levels, reductions in clutch andlitter size, and alterations in reproductive behavior), andthe mechanisms believed to cause them. We believe these arethe direct toxic effects most likely to reduce populations offree-living birds and mammals within treated areas. Data fromstudies of captive birds and laboratory mammals and free-livingindividuals given controlled dosages are included with an emphasison information published within the last 5 years. The limitationsof existing data for determining the biological and regulatorysignificance of these effects are discussed.     

Pigmented Human Skin Equivalent: New Method of Reconstitution by Grafting an Epithelial Sheet Onto a Non-Contractile Dermal Equivalent

We have established a new protocol for reconstituting a pigmented human skin equivalent (PSE) and have evaluated its functional responses to environmental stimulus, UVB. The PSE is reconstituted by grafting an epithelial sheet consisting of keratinocytes and melanocytes onto a porous non-contractile dermal equivalent populated with mitotically and metabolically active fibroblasts. i) The PSE has a multilayered, well-differentiated epidermis with cuboidal basal cells and highly organised dermis with newly synthesised extracellular matrix components. ii) Ki67-positive proliferating keratinocytes (18.1 ± 7.4%) were detected on the basal layer of the epidermis. iii) Melanocytes located exclusively within the basal layer were detected by monoclonal antibody against tyrosinase-related protein (TRP-1). iv) After exposure to UVB (100 mJ/cm² per day) for 7 consecutive days, the intensity of TRP-1 staining was increased in the PSE, showing their functional state, whereas the number of melanocytes was not changed. This non-contractile and functioning new PSE is potentially useful as a model for studying the role of melanocyte-keratinocyte-fibroblast interactions in photoprotection of the skin in more complex cutaneous microenvironment than monolayer culture, and for developing in vitro disease models and therapeutic protocols with genetically altered cells both in epidermis and dermis.     

Monoterpenes emitted by the large pine weevil, Hylobius abietis (L.) feeding on Scots pine, Pinus sylvestris L.

Abstract. Attraction of the large pine weevil, Hylobius abietis (L.) (Coleoptera: Curculionidae) to pine logs was enhanced by the presence of weevils already on the logs. We investigated the volatiles emitted by insects feeding on host trees to determine the mechanism of the attraction. Chemical analyses of the collected volatiles showed large amounts of monoterpenes. There were no qualitative differences in the collections, between Scots pine alone and pine plus feeding weevils, nor between males and females, mated or virgin. Strong quantitative differences in monoterpenes existed that were correlated to the surface of cut bark. Electroantennograms (EAGs) were recorded on male and female antennae stimulated by increasing doses of collected volatiles. EAGs for pine alone and pine plus weevils at the same stimulus loading in (a + P)-pinene gave superimposable curves. EAGs of male and female volatiles were also identical. These results suggest an attraction mediated by host plant allelochemicals rather than by an aggregation pheromone.