Polarized THG Microscopy Identifies Compositionally Different Lipid Droplets in Mammalian Cells

Cells store excess lipids as two major compounds, triacylglycerols (TAGs) and cholesteryl esters (CEs), inside lipid droplets (LDs). The degree of lipid ordering is considered to play a major role in the mobility and enzymatic processing of lipids in LDs. Here, we provide evidence that polarized third-harmonic generation (THG) microscopy distinguishes between native TAG- and CE-enriched LDs in cells due to the different ordering of the two lipid species. We first demonstrate that the responses from synthetic TAG- and CE-enriched LDs using THG microscopy with linear and circular polarizations differ according to their different intrinsic ordering. We then employ simulations to dissect how polarization effects influence the THG from an isotropic LD. Finally, we induce TAG- and CE-enriched LDs in murine macrophages and demonstrate that polarized THG responses increase in a nonlinear fashion with increasing CE/TAG ratio. This suggests that with an increasing CE content, there is a rather sharp transition toward increased LD ordering. Our results demonstrate that polarized THG microscopy enables label-free quantitative analysis of LD ordering and discriminates between compositionally different LDs in intact mammalian cells.     

Sexual incentive motivation in male rats requires both androgens and estrogens

In Experiment 1 castrated male rats were implanted with a Silastic capsule containing either E or cholesterol (CHOL) 35 days after castration. They were then tested for sexual incentive motivation and copulatory behaviors every 5th day for 3 weeks. None of the treatments affected sexual incentive motivation. After the last test, all subjects were implanted with DHT-containing Silastic capsules, and tests continued for another 3 weeks. While E + DHT enhanced sexual incentive motivation and copulatory behavior, DHT alone failed to do so. In Experiment 2 the aromatase inhibitor fadrozole (F) was combined with testosterone (T). T restored all behaviors to the level seen in intact rats, and F significantly reduced these effects. In fact, T + F was not different from DHT. T and DHT restored the weight of the prostate and seminal vesicles to levels close to those of intact rats. In Experiment 3 a lower dose of E was employed. Also this dose of E failed to affect sexual incentive motivation while E + DHT restored it to the level of intact animals. Castration enhanced the serum concentrations of LH and FSH. E alone caused a marked reduction, and E + DHT brought both gonadotropins back to the level of intact animals. It was concluded that the doses of E and DHT employed in these experiments were within or close to the physiological range, and that such doses of E completely fail to enhance sexual incentive motivation in castrated animals. DHT has small or no effects. It appears that sexual incentive motivation and copulation require simultaneous stimulation of androgen and estrogen receptors.     

Effects of group size and pine defence chemicals on Diprionid sawfly survival against ant predation

The defence chemicals and behavioural adaptations (gregariousness and active defensive behaviour) of pine sawfly larvae may be effective against ant predation. However, previous studies have tested their defences against very few species of ants, and few experiments have explored ant predation in nature. We studied how larval group size (groups of 5 and 20 in Neodiprion sertifer and 10, 20 and 40 in Diprion pini) and variation in levels of defence chemicals in the host tree (Scots pine, Pinus sylvestris) affect the survival of sawfly larvae. Food preference experiments showed that ants do eat sawfly larvae, although they are not their most preferred food item. According to our results, ant predation significantly increases the mortality rate of sawfly larvae. Larval mortality was minor on pine tree branches where ant traffic was excluded. We also found that a high resin acid concentration in the host tree significantly decreased the mortality of D. pini larvae when ants were present. However, there was no such relationship between the chemical concentrations of the host tree and larval mortality for N. sertifer. Surprisingly, grouping did not help sawfly larvae against ant predation. Mortality risk was the same for all group sizes. The results of the study seemingly contradict previous understanding of the effectiveness of defence mechanisms of pine sawfly against ant predation, and suggest that ants (Formica exsecta in particular) are effective predators of sawfly larvae.An erratum to this article can be found at     

A 3-year exposure to CO2 and O3 induced minor changes in soil N cycling in a meadow ecosystem

We investigated the individual and interactive effects of moderately elevated CO₂ (ambient air + 100 ppm) and/or O₃ (40–50 ppb) on soil N cycling and microbial biomass N in a 3-year open-top chamber experiment conducted in meadow mesocosms. The results show that elevated O₃ decreased the concentrations of mineral N and NH₄⁺-N in the mesocosm soil in the last growing season (2004). Total N, NO₃⁻-N, microbial biomass N, decomposition rate, potential nitrification and denitrification were not affected by elevated O₃ and/or CO₂. It is thus concluded that the proposed future ambient O₃ and CO₂ levels, such as used in this experiment, may not induce major changes in the below-ground N processes in N-poor northern European hay meadow ecosystems.     

Impacts of climate warming on the long-term dynamics of key fish species in 24 European lakes

Fish play a key role in the trophic dynamics of lakes. With climate warming, complex changes in fish assemblage structure may be expected owing to direct effects of temperature and indirect effects operating through eutrophication, water level changes, stratification and salinisation. We reviewed published and new long-term (10–100 years) fish data series from 24 European lakes (area: 0.04–5,648 km²; mean depth: 1–177 m; a north–south gradient from Sweden to Spain). Along with an annual temperature increase of about 0.15–0.3°C per decade profound changes have occurred in either fish assemblage composition, body size and/or age structure during recent decades and a shift towards higher dominance of eurythermal species. These shifts have occurred despite a reduction in nutrient loading in many of the lakes that should have benefited the larger-sized individuals and the fish species typically inhabiting cold-water, low-nutrient lakes. The cold-stenothermic Arctic charr has been particularly affected and its abundance has decreased in the majority of the lakes where its presence was recorded. The harvest of cool-stenothermal brown trout has decreased substantially in two southern lakes. Vendace, whitefish and smelt show a different response depending on lake depth and latitude. Perch has apparently been stimulated in the north, with stronger year classes in warm years, but its abundance has declined in the southern Lake Maggiore, Italy. Where introduced, roach seems to take advantage of the higher temperature after years of low population densities. Eurythermal species such as common bream, pike–perch and/or shad are apparently on the increase in several of the lakes. The response of fish to the warming has been surprisingly strong and fast in recent decades, making them ideal sentinels for detecting and documenting climate-induced modifications of freshwater ecosystems.     

Assessment of the ecological status of regulated lakes: stressor-specific metrics from littoral fish assemblages

Although in situ sediment capping is frequently used to reduce internal loading of contaminants and nutrients, post-application assessment rarely includes the potential undesirable short-term effects on plankton species composition. We hypothesised that a modified zeolite (Z2G1) application as a sediment capping agent in Lake Okaro, New Zealand, could cause significant undesirable shifts in species composition of both zooplankton and phytoplankton due to burial of resting stages or interference with feeding for the zooplankton. Alternatively, we predicted that the capping agent might have no effect due to, for example, the coarse grain size of the material (1–3 mm). We used multidimensional scaling (MDS) and analysis of similarity (ANOSIM) to identify any adverse effects of Z2G1 on zooplankton and phytoplankton species composition (i.e. shifts in community structure, including species loss) by comparing the community structure before and after the Z2G1 application. We found no significant differences in species composition before and after the Z2G1 application at the depths investigated (surface and 9 m). However, all of the analyses showed statistically significant differences among seasons, indicating seasonal variations in plankton composition far outweigh those that may have resulted from the Z2G1 application. Coarse particle size, low dose rate and a restricted area where the sediment capping agent was applied were considered to be the factors limiting potential adverse effects on plankton species. Considerations of finer-grained material to increase coverage and efficacy of phosphorus adsorption require assessment for their effects on zooplankton, however, and a direct mode of application into the hypolimnion is recommended to minimise effects on zooplankton and phytoplankton communities.     

Shiga toxin-producing Escherichia coli O100:H⁻: stx2e in drinking water contaminated by waste water in Finland

In November 2007, 450 m³ of treated wastewater leaked into the drinking water distribution system contaminating the drinking water of over 10,000 inhabitants of Nokia, Southern Finland. Nearly 1,000 people visited the health centre because of gastroenteritis during the following 5 weeks. A wide range of enteric pathogens was found in the patients. The authors used the 16-plex PCR to investigate whether the five major diarrheagenic Escherichia coli pathotypes (EPEC, ETEC, STEC, EIEC or EAEC) were present in the contaminated drinking water and in the patients’ stool samples. The contaminated drinking water was positive for genes characteristic of various E. coli pathotypes: pic, invE, hlyA, ent, escV, eae, aggR, stx ₂ , estIa and astA. These genes, except stx ₂ , hlyA and invE, were also detected in the stool samples of the patients linked to this outbreak. A sorbitol positive, streptomycin resistant STEC strain was isolated from the drinking water, and belonged to the serotype O100:H ^– , produced Stx2 toxin (titre 1:8 by reversed-passive latex agglutination method), and carried the genes stx _2e, estIa and irp2.     

Novel Microbiological and Spatial Statistical Methods to Improve Strength of Epidemiological Evidence in a Community-Wide Waterborne Outbreak

Failures in the drinking water distribution system cause gastrointestinal outbreaks with multiple pathogens. A water distribution pipe breakage caused a community-wide waterborne outbreak in Vuorela, Finland, July 2012. We investigated this outbreak with advanced epidemiological and microbiological methods. A total of 473/2931 inhabitants (16%) responded to a web-based questionnaire. Water and patient samples were subjected to analysis of multiple microbial targets, molecular typing and microbial community analysis. Spatial analysis on the water distribution network was done and we applied a spatial logistic regression model. The course of the illness was mild. Drinking untreated tap water from the defined outbreak area was significantly associated with illness (RR 5.6, 95% CI 1.9–16.4) increasing in a dose response manner. The closer a person lived to the water distribution breakage point, the higher the risk of becoming ill. Sapovirus, enterovirus, single Campylobacter jejuni and EHEC O157:H7 findings as well as virulence genes for EPEC, EAEC and EHEC pathogroups were detected by molecular or culture methods from the faecal samples of the patients. EPEC, EAEC and EHEC virulence genes and faecal indicator bacteria were also detected in water samples. Microbial community sequencing of contaminated tap water revealed abundance of Arcobacter species. The polyphasic approach improved the understanding of the source of the infections, and aided to define the extent and magnitude of this outbreak.     

Effects of polarization in the presence and absence of biocides on biofilms in a simulated paper machine water

The antifouling potential of electric polarization combined and not combined with biocides was studied in nonsaline warm water with high organic content. Deinococcus geothermalis is a bacterium known for forming colored biofilms in paper machines and for its persistence against cleaning and chemical treatments. When D. geothermalis biofilms grown for 24 h in simulated paper machine water were exposed to cathodic or cathodically weighted pulsed polarization at least 60% (P < 0.05) of the biofilms were removed from stainless steel (AISI 316L). Biofilm removal by 25 ppm (effective substances 5-25 ppm) of oxidizing biocides (bromochloro-5,5-dimethylhydantoin, 2,2-dibromo-2-cyanoacetamide, peracetic acid) increased to 70% when combined with cathodically weighted pulsed polarization. Using a novel instrument that allows real-time detection of reactive oxygen species (ROS) we showed that the polarization program effective in antifouling generated ROS in a pulsed manner on the steel surface. We thus suggest that the observed added value of oxidative biocides combined with polarization depended on ROS. This suggestion was supported by the finding that a reductive biocide, methylene bisthiocyanate, counteracted the antifouling effect of polarization.