Characterization of <Emphasis Type="Italic">fs10.1</Emphasis>, a major QTL controlling fruit elongation in <Emphasis Type="Italic">Capsicum</Emphasis>

We previously identified fs10.1 as a major QTL controlling fruit shape (index of length to width) in an interspecific F₂ cross of Capsicum annuum (round fruit) × C. chinense (elongated fruit) in pepper. To more precisely map and characterize the QTL, we constructed near-isogenic lines for fs10.1 and mapped it in a BC₄F₂ population. In this population, fs10.1 segregated as a Mendelian locus and mapped 0.3 cM away from the closest molecular marker. We further verified the effect of fs10.1 in an F₂ population from an independent cross between elongated- and conical-fruited parents. To identify additional allelic variation at fruit shape loci, we screened an EMS-mutagenized population of the blocky-fruited cv. Maor and identified the mutant E-1654 with elongated fruit. This fruit shape mutation was mapped to the fs10.1 region and was determined to be allelic to the QTL. By measuring fruit shape of near-isogenic lines for fs10.1 during fruit development, we found that the shape of the fruit is determined primarily in the first 2 weeks after anthesis. Histological measurements of cell size and cell shape in pericarp sections of fruits of the isogenic lines throughout fruit development indicated that the shape of the fruit is determined primarily by cell shape and that the development of fruit shape is correlated with cell shape.     

Genome sequence analyses of two isolates from the recent <Emphasis Type="Italic">Escherichia coli</Emphasis> outbreak in Germany reveal the emergence of a new pathotype: Entero-Aggregative-Haemorrhagic <Emphasis Type="Italic">Escherichia coli</Emphasis> (EAHEC)

The genome sequences of two Escherichia coli O104:H4 strains derived from two different patients of the 2011 German E. coli outbreak were determined. The two analyzed strains were designated E. coli GOS1 and GOS2 (German outbreak strain). Both isolates comprise one chromosome of approximately 5.31 Mbp and two putative plasmids. Comparisons of the 5,217 (GOS1) and 5,224 (GOS2) predicted protein-encoding genes with various E. coli strains, and a multilocus sequence typing analysis revealed that the isolates were most similar to the entero-aggregative E. coli (EAEC) strain 55989. In addition, one of the putative plasmids of the outbreak strain is similar to pAA-type plasmids of EAEC strains, which contain aggregative adhesion fimbrial operons. The second putative plasmid harbors genes for extended-spectrum β-lactamases. This type of plasmid is widely distributed in pathogenic E. coli strains. A significant difference of the E. coli GOS1 and GOS2 genomes to those of EAEC strains is the presence of a prophage encoding the Shiga toxin, which is characteristic for enterohemorrhagic E. coli (EHEC) strains. The unique combination of genomic features of the German outbreak strain, containing characteristics from pathotypes EAEC and EHEC, suggested that it represents a new pathotype Entero-Aggregative-Haemorrhagic E scherichia c oli (EAHEC).     

Δ<Superscript>6</Superscript>-Desaturase from <Emphasis Type="Italic">Mortierella alpina</Emphasis>: cDNA cloning,expression, and phylogenetic analysis

The open reading frame of the Δ⁶-desaturase gene was isolated from Mortierella alpina W15 and the gene was cloned into a pPIC3.5K vector. The vector was transformed into Pichia pastoris GS115 and expression was induced with methanol. The Δ⁶-desaturase expressed in P. pastoris GS115 catalyzed the conversion of linoleic acid to γ-linolenic acid but not the conversion of α-linolenic acid to octadecatetraenoic acid. The results indicate that the Δ⁶-desaturase gene from M. alpina W15 has substrate specificity in different organisms. Phylogenetic analysis revealed that Δ⁶-desaturase genes can be divided into four monophyletic groups. This work paves the way for further study of the functions of Δ⁶-desaturase in fatty acid metabolism and its three-dimensional structure.     

The Effect of Sterilization Methods on the Physical Properties of Silk Sericin Scaffolds

Protein-based biomaterials respond differently to sterilization methods. Since protein is a complex structure, heat, or irradiation may result in the loss of its physical or biological properties. Recent investigations have shown that sericin, a degumming silk protein, can be successfully formed into a 3-D scaffolds after mixing with other polymers which can be applied in skin tissue engineering. The objective of this study was to investigate the effectiveness of ethanol, ethylene oxide (EtO) and gamma irradiation on the sterilization of sericin scaffolds. The influence of these sterilization methods on the physical properties such as pore size, scaffold dimensions, swelling and mechanical properties, as well as the amount of sericin released from sericin/polyvinyl alcohol/glycerin scaffolds, were also investigated. Ethanol treatment was ineffective for sericin scaffold sterilization whereas gamma irradiation was the most effective technique for scaffold sterilization. Moreover, ethanol also caused significant changes in pore size resulting from shrinkage of the scaffold. Gamma-irradiated samples exhibited the highest swelling property, but they also lost the greatest amount of weight after immersion for 24 h compared with scaffolds obtained from other sterilization methods. The results of the maximum stress test and Young’s modulus showed that gamma-irradiated and ethanol-treated scaffolds are more flexible than the EtO-treated and untreated scaffolds. The amount of sericin released, which was related to its collagen promoting effect, was highest from the gamma-irradiated scaffold. The results of this study indicate that gamma irradiation should have the greatest potential for sterilizing sericin scaffolds for skin tissue engineering.     

Molecular Characterization of Fecal Microbiota in Patients with Viral Diarrhea

The study provides molecular analyses of fecal microbiota of diarrhea patients infected with four different types of viruses. Fecal specimens from 52 patients with viral diarrhea (13 each of adenovirus, norovirus, rotavirus, and astrovirus) and six healthy individuals were collected and etiological viral agent was confirmed by enzyme immunoassay and specific PCR. To assess the changes in microbial diversity in patients with viral diarrhea, DNA from stool were extracted and characterized by PCR-denaturing gradient gel electrophoresis (DGGE) with universal primers specific for the V3 region of 16S rRNA gene. The strongest bands of the DGGE profiling were excised and sequenced to identify the dominant groups. Bacteroides vulgatus, Bifidobacterium, and Lactobacillus genera were also enumerated by real time PCR. The results revealed that bacterial diversity and similarity in feces from viral diarrhea groups were significantly lower (mean H′/ H _max^￠ H_{ \max }^{\prime } 0.89–0.94, 29–43, respectively) as compared with those of healthy individuals (mean H′/ H _max^￠ H_{ \max }^{\prime } 1.36, 59, respectively). Sequencing of dominant bands affirmed that diarrhea groups were mainly comprised of phylum Firmicutes, such as genera Enterococcus, Peptostreptococcaceae incertae sedi, Streptococcus, Weissella, and Clostridium, and opportunistically pathogenic genus Shigella, while dominant group in healthy individuals was phylum Bacteroidetes. Copy number of Bacteroides vulgatus, Bifidobacterium, and Lactobacillus genera was also reduced significantly in viral diarrhea groups as compared to healthy group. It is concluded that opportunistic pathogens increases, while other species of commensal microbiota decrease significantly in the viral diarrhea patients and dysbacteriosis is dependent on type of virus infection.     

Effects of low temperature on growth and non-structural carbohydrates of the imperial bromeliad <Emphasis Type="Italic">Alcantarea imperialis</Emphasis> cultured in vitro

The imperial bromeliad Alcantarea imperialis grows naturally on rocky outcrops (‘inselbergs’) in regions where daily temperatures vary from 5 to 40°C. As carbohydrate metabolism is altered in response to cold, it could lead to reprogramming of the metabolic machinery including the increase in levels of metabolites that function as osmolytes, compatible solutes, or energy sources in order to maintain plant homeostasis. The aim of this study was to evaluate the effects of different temperatures on plant growth and non-structural carbohydrates in plants of A. imperialis adapted to low temperature. Seedlings of A. imperialis were grown in vitro under a 12-h photoperiod with four different day/night temperature cycles: 5/5°C, 15/15°C, 15/30°C (dark/light) and 30/30°C. Plants were also cultivated at 26°C in ex vitro conditions for comparison. The results showed an inverse relationship between temperature and germination time and no differences in the percentage of germination. Plants maintained for 9 months at 15°C presented a reduced number of leaves and roots, and a dry mass four times lower than plants grown at 30°C. Sugar content was higher in plants grown at 15°C than at 30°C. However, the highest amount of total sugar was found in plants growing under warm day/cold night conditions. Myo-inositol, glucose, fructose and sucrose were found predominantly under high temperatures, while under low temperatures, sucrose was apparently replaced by trehalose, raffinose and stachyose. Starch content was highest in plants grown under high temperatures. The lowest starch content was detected under low temperatures, suggesting its conversion into soluble carbohydrates to protect the plants against cold. These results indicated that low temperature retarded growth of A. imperialis and increased sugar levels, mainly trehalose, thus suggesting that these sugar compounds could be involved in cold tolerance.     

Novel three-phase bioreactor concept for fatty acid alkyl ester production using <Emphasis Type="Italic">R. oryzae</Emphasis> as whole cell catalyst

A novel three-phase solid–gas–liquid bioreactor (SGLB) concept using gaseous alcohol and liquid rapeseed oil with immersed microorganisms overlying a nutrient agar phase (solid) is proposed for biodiesel (fatty acid alkyl esters, FAAE) production based on the high hydrophobicity and negative surface charge showed by the fungi Rhizopus oryzae. This novel bioreactor was thought to increase oil bioavailability and decrease alcohol toxicity for effective microbial growth, reaching high yields of FAAE production without any pretreatment. High growth rates were reached for R. oryzae using a SGLB simultaneously reaching a high FAAE production yield, up to 50% using methanol and up to 70% using ethanol at 144 h of incubation at 20°C. To compare the effect of gaseous alcohol, the same experiments were carried out in a three-phase solid–liquid–liquid bioreactor (SLLB), where the alcohol was added in liquid phase, showing significant R. oryzae growth but no FAAE formation. This suggests that the inhibitory effect of alcohol is more significant in lipase activity than in R. oryzae growth, and the use of alcohol in gaseous phase may decrease both of them. The experimental procedure using SGLB showed that when R. oryzae is maintained alive, it can catalyze at the same time the hydrolysis, esterification and transesterification of triglycerides from rapeseed oil, but its activity strongly depends on the used growth media. Therefore, the application of gaseous alcohol coupled with R. oryzae as immobilized whole cell catalysts may be a potential alternative to the use of commercial lipases for biodiesel production.     

Additive effects of herbivory,nectar robbing and seed predation on male and female fitness estimates of the host plant <Emphasis Type="Italic">Ipomopsis aggregata</Emphasis>

Many antagonistic species attack plants and consume specific plant parts. Understanding how these antagonists affect plant fitness individually and in combination is an important research focus in ecology and evolution. We examined the individual and combined effects of herbivory, nectar robbing, and pre-dispersal seed predation on male and female estimates of fitness in the host plant Ipomopsis aggregata. By examining the effects of antagonists on plant traits, we were able to tease apart the direct consumptive effects of antagonists versus the indirect effects mediated through changes in traits important to pollination. In a three-way factorial field experiment, we manipulated herbivory, nectar robbing, and seed predation. Herbivory and seed predation reduced some male and female fitness estimates, whereas plants tolerated the effects of robbing. The effects of herbivory, robbing, and seed predation were primarily additive, and we found little evidence for non-additive effects of multiple antagonists on plant reproduction. Herbivory affected plant reproduction through both direct consumptive effects and indirectly through changes in traits important to pollination (i.e., nectar and phenological traits). Conversely, seed predators primarily had direct consumptive effects on plants. Our results suggest that the effects of multiple antagonists on estimates of plant fitness can be additive, and investigating which traits respond to damage can provide insight into how antagonists shape plant performance.     

Alkaline lipase activity from the marine protists,thraustochytrids

Two thraustochytrid protists of the genus Thraustochytrium isolated from coastal and mangrove habitats of Goa, India were studied for extracellular alkaline lipase production. Maximum lipase production was supported by a combination of peptone and yeast extract in the growth medium while strong inhibition of enzyme production was observed in presence of glucose. The inducible nature of the enzyme production was evidenced by the requirement of olive oil in the medium. Lipase production was salt-dependent and optimum production required 3.4% (w/v) crude sea salt. Ideal conditions for maximum production of lipases were therefore adopted as incubation at 30 ± 2°C for 168 h at an initial pH of 6.0 in a medium consisting of 0.5% peptone, 0.01% yeast extract, 0.5% olive oil and 3.4% crude salt. Extracellular lipase production by the two thraustochytrid isolates [designated TZ (ATCC #PRA-295) and AH-2 (ATCC #PRA-296)] was increased threefold under these optimized culture conditions. This appears to be the first report on optimization of cultivation conditions for the production of alkaline lipases by thraustochytrids.