Gene Expression Analysis of Cold and Freeze Stress in Baker's Yeast

We used mRNA differential display to assess yeast gene expression under cold or freeze shock stress conditions. We found both up- and down-regulation of genes, although repression was more common. We identified and sequenced several cold-induced genes exhibiting the largest differences. We confirmed, by Northern blotting, the specificity of the response for TPI1, which encodes triose-phosphate isomerase; ERG10, the gene for acetoacetyl coenzyme A thiolase; and IMH1, which encodes a protein implicated in protein transport. These genes also were induced under other stress conditions, suggesting that this cold response is mediated by a general stress mechanism. We determined the physiological significance of the cold-induced expression change of these genes in two baker's yeast strains with different sensitivities to freeze stress. The mRNA level of TPI1 and ERG10 genes was higher in freeze-stressed than in control samples of the tolerant strain. In contrast, both genes were repressed in frozen cells of the sensitive strain. Next, we examined the effects of ERG10 overexpression on cold and freeze-thaw tolerance. Growth of wild-type cells at 10°C was not affected by high ERG10 expression. However, YEpERG10 transformant cells exhibited increased freezing tolerance. Consistent with this, cells of an erg10 mutant strain showed a clear phenotype of cold and freeze sensitivity. These results give support to the idea that a cause-and-effect relationship between differentially expressed genes and cryoresistance exists in Saccharomyces cerevisiae and open up the possibility of design strategies to improve the freeze tolerance of baker's yeast.     

Isolation and PCR detection of <Emphasis Type="Italic">Enterobacter sakazakii</Emphasis> in South African food products,specifically infant formula milks

Enterobacter sakazakii has recently been identified as an opportunistic pathogen. The current culture-dependent detection methods for these bacteria are time-consuming and in this study a PCR method for the detection of E. sakazakii in South African food products, including an internal amplification control (IAC) was developed. DNA was isolated and amplified from the products and they were plated on selective growth media after pre-enrichment and enrichment in Enterobacteriaceae enrichment broth. Four of the 22 products tested positive for the presence of E. sakazakii, confirmed by PCR detection and growth on selective media. The PCR method proved effective in detecting E. sakazakii in South African products after three days and could serve as an alternative for traditional microbiological techniques.     

Rock selection for nesting inProformica longiseta Collingwood, 1978 (Hymenoptera,Formicidae) in a Mediterranean high mountain

Proformica longiseta, an ant species endemic to certain high mountains, has been studied with regard to its tendency to nest underneath rocks. This behaviour led us to investigate whether a particular type of rock is selected, and, if so, the advantages to this species of the selection. This study was located at 2400 m above sea-level in the Sierra Nevada mountains (Granada, Spain). To determine the range of rock sizes available, in eight control plots the thicknesses were recorded and also the maximum and minimum diameters of all rocks with surface area of more than 6 cm². A total of 1724 rocks were recorded, noting among the rock types available those that were used for nesting. Also as a preliminary estimate of the thermal characteristics of the rocks, we recorded the temperatures of the air, soil surface, soil under a rock covering an ant nest, and soil at 30 cm depth. The results indicate thatP. longiseta selects flat rocks with a surface area/thickness ratio of 2–5, the thickness being not less than 1.5 cm, the surface area being 50–250 cm² and volume less than 1000 cm³. This type of rock offers the nests two beneficial but apparently contradictory effects: (1) protection against high temperatures on hot days, and (2) warmth on colder days by absorbing the diffusing solar heat more effectively than the air or soil around the nest.     

The hypotensive drug 2-hydroxyoleic acid modifies the structural properties of model membranes

We studied the interactions of the hypotensive drug, 2-hydroxyoleic acid (2OHOA), with model membranes using the techniques of DSC, ³¹P NMR and X-ray diffraction. We demonstrate that 2OHOA alters the thermotropic behaviour of 1,2-dielaidoyl-sn-glycero-3-phosphoethanolamine (DEPE), thereby promoting the formation of hexagonal phases (H_II), despite stabilizing the lamellar phase (L_α). The lattice parameters of lamellar and non-lamellar structures were not altered by the presence of 2OHOA. The molecular bases underlying the alterations in membrane structure provoked by 2OHOA were analysed by comparing the effects produced by 2OHOA with the closely related fatty acids (FAs), oleic acid (OA) and elaidic acid (EA). The capacity of C-18 FAs to induce H_II-phase formation followed the order OA>2OHOA>EA. Furthermore, while 2OHOA stabilized the L_α phase, OA destabilized it. The net negative charge of 2OHOA at physiological pH (～7.4) influenced its effect on membrane structure. By analysing the molecular architecture of 2OHOA in DEPE monolayers, interactions between the carboxylate groups of 2OHOA and the amine groups of DEPE were observed, as well as between the 2-hydroxyl group of the FA and the carbonyl oxygen of the phospholipid acyl chain. These structural characteristics provoked an increase in the P-to-N and P-to-P distances of neighbouring phospholipid headgroups in the presence of 2OHOA, with respect to those observed with OA and EA. The higher headgroup area at the lipid–water interface in presence of 2OHOA could account for the differential effect of this drug on the phase behaviour of DEPE membranes.     

High-Throughput 3-D Monitoring of Agricultural-Tree Plantations with Unmanned Aerial Vehicle (UAV) Technology

The geometric features of agricultural trees such as canopy area, tree height and crown volume provide useful information about plantation status and crop production. However, these variables are mostly estimated after a time-consuming and hard field work and applying equations that treat the trees as geometric solids, which produce inconsistent results. As an alternative, this work presents an innovative procedure for computing the 3-dimensional geometric features of individual trees and tree-rows by applying two consecutive phases: 1) generation of Digital Surface Models with Unmanned Aerial Vehicle (UAV) technology and 2) use of object-based image analysis techniques. Our UAV-based procedure produced successful results both in single-tree and in tree-row plantations, reporting up to 97% accuracy on area quantification and minimal deviations compared to in-field estimations of tree heights and crown volumes. The maps generated could be used to understand the linkages between tree grown and field-related factors or to optimize crop management operations in the context of precision agriculture with relevant agro-environmental implications.     

Co-fractionation of an endonuclease activity during the purification of DNA polymerase-alpha from regenerating rat liver. Properties and separation from DNA polymerase.

The presence of endonuclease activity associated with DNA polymerase was detected during the purification of high-molecular-weight DNA polymerase-alpha from regenerating rat liver by the use of a highly sensitive test. This endonuclease activity co-fractionated with DNA polymerase in a great variety of purification procedures involving ion-exchange chromatographies or molecular weight fractionation, but was further completely separated from DNA polymerase activity by using affinity chromatography on DNA-cellulose. The endonuclease acted on native or denatured DNA by introducing single-strand nicks in the DNA molecules; its enzymatic properties indicate that it could act in polymerisation conditions in vitro.     

Effects of habitat simplification on assemblages of cavity nesting bees and wasps in a semiarid neotropical conservation area

Habitat complexity is directly correlated to insect diversity in most natural environments. Structural complexity reflects an increase in vertical stratification and plant diversity and often leads to a greater availability of floral resources and nesting sites. Efficient conservation strategies require understanding of how changes in habitat structure affect insects that provide essential ecosystem services. We analyzed how the diversity and species composition of bees and wasps that nest in pre-existing cavities is affected by habitat complexity. Our study was developed in the semiarid region of northeastern Brazil, in the Ubajara National Park and surrounding area. Four types of habitats within two physiognomies were sampled for two consecutive years. We used 120 trap-nest (9000 cavities) distributed in 40 sample points. Overall, 657 cavities were occupied by 11 species of bees, nine of wasps, and six of cleptoparasitic/parasitoids. Bees and wasp diversity increases with habitat complexity. While species richness was higher in more complex physiognomies, abundance was higher in disturbed areas. Species composition also varied with habitat structure. Habitat simplification has adverse effects on the diversity and composition of assemblages. These effects are stronger in more complex habitats indicating that conservation of humid habitats within semiarid areas is essential to maintain bee and wasp regional diversity.