Site-specific mutations in the D1 polypeptide affect the susceptibility of Synechocystis 6803 cells to photoinhibition

Photoinhibition of photosystem II in the cyanobacterium Synechocystis 6803 was followed after site-specific mutagenesis of the D1 polypeptide. Mutations were created in the stromal/cytosolic loop connecting helices D and E. Two mutations E243K and CA1, a deletion of the three glutamates 242–244 and a substitution Q241H, were made in the putative cleavage area of the D1 polypeptide. A third mutation E229D was made in the PEST-like sequence. Mutants and control cells were illuminated and F_V/F_M was recorded. Compared to the control, the mutants were less photoinhibited. Fluorescence relaxation after a single flash was delayed in CA1. Restoration of F_V/F_M after photoinhibition in the mutants was totally dependent on protein synthesis while control cells were able to recover partially also when protein synthesis was inhibited. In addition, the protein synthesis-dependent recovery of CA1 was slowed down. Our results indicate a correlation between the mutated amino acids and photoinhibition of photosystem II.     

High frequency one-step gene replacement in Trichoderma reesei. II. Effects of deletions of individual cellulase genes

Four cellulase genes of Trichoderma reesei, cbh1, cbh2, egl1 and egl2, have been replaced by the amdS marker gene. When linear DNA fragments and flanking regions of the corresponding cellulase locus of more than 1 kb were used, the replacement frequencies were high, ranging from 32 to 52%. Deletion of the major cellobiohydrolase 1 gene led to a 2-fold increase in the production of cellobiohydrolase II; however, replacement of the cbh2 gene did not affect the final cellulase levels and deletion of egl1 or egl2, slightly increased production of both cellobiohydrolases. Based on our results, endoglucanase II accounts for most of the endoglucanase activity produced by the hypercellulolytic host strain. Furthermore, loss of the egl2, gene causes a significant drop in the filter paper-hydrolysing activity, indicating that endoglucanase II has an important role in the total hydrolysis of cellulose.     

High frequency one-step gene replacement in Trichoderma reesei. II. Effects of deletions of individual cellulase genes

Four cellulase genes of Trichoderma reesei, cbh1, cbh2, egl1 and egl2, have been replaced by the amdS marker gene. When linear DNA fragments and flanking regions of the corresponding cellulase locus of more than 1 kb were used, the replacement frequencies were high, ranging from 32 to 52%. Deletion of the major cellobiohydrolase 1 gene led to a 2-fold increase in the production of cellobiohydrolase II; however, replacement of the cbh2 gene did not affect the final cellulase levels and deletion of egl1 or egl2, slightly increased production of both cellobiohydrolases. Based on our results, endoglucanase II accounts for most of the endoglucanase activity produced by the hypercellulolytic host strain. Furthermore, loss of the egl2, gene causes a significant drop in the filter paper-hydrolysing activity, indicating that endoglucanase II has an important role in the total hydrolysis of cellulose.     

Changes of amino acid sequence in PEST-like area and QEEET motif affect degradation rate of D1 polypeptide in photosystem II

The degradation rate of the D1 polypeptide was measured in threeSynechocystis PCC 6803 mutantsin vivo. Mutations were introduced into a putative cleavage area of the D1 polypeptide (QEEET motif) and into the PEST-like area. PEST sequences are often found in proteins with a high turnover rate. The QEEET-motif mutants are CA1 [(E242-E244);Q241H] and E243K, and the third mutation, E229D, was directed to the PEST-like area. During high-light illumination (1500 mol photons m^-2s^-1) that induced photoinhibition of photosystem II (PSII), the half-life time of the D1 polypeptide in mutant E229D (t _1/2=35 min) was about twice as long as in AR (control strain) cells (t _1/2=19 min). In growth light (40 mol photons m^-2s^-1), the degradation rate of the D1 polypeptide in E229D and AR strains was the same (t _1/25 h). In growth light the D1 polypeptide was degraded faster in both QEEET-motif mutants than in the AR strain, but in photoinhibitory light the degradation rates were similar. According to these results, the highly conservative QEEET motif as such is not required for the proteolytic cut of the D1 polypeptide, but it does affect the rate of degradation. No simple correlation existed between the degradation rate of the D1 polypeptide and the susceptibility of PSII to photoinhibition in mutant and AR cells under our experimental conditions.     

Maximum Temperature Limits for Acidophilic, Mesophilic Bacteria in Biological Leaching Systems

The maximum temperature for growth (T_max) was determined for pure and mixed cultures of acidophilic thiobacilli. The experimental system was based on incubating the cultures in liquid media exposed to a linear temperature gradient. The T_max values varied within the range of 36.1 to 43.6°C.     

Harsh nutritional environment has positive and negative consequences for family living in a burying beetle

Harsh environmental conditions in form of low food availability for both offspring and parents alike can affect breeding behavior and success. There has been evidence that food scarce environments can induce competition between family members, and this might be intensified when parents are caring as a pair and not alone. On the other hand, it is possible that a harsh, food-poor environment could also promote cooperative behaviors within a family, leading, for example, to a higher breeding success of pairs than of single parents. We studied the influence of a harsh nutritional environment on the fitness outcome of family living in the burying beetle Nicrophorus vespilloides. These beetles use vertebrate carcasses for reproduction. We manipulated food availability on two levels: before and during breeding. We then compared the effect of these manipulations in broods with either single females or biparentally breeding males and females. We show that pairs of beetles that experienced a food-poor environment before breeding consumed a higher quantity of the carcass than well-fed pairs or single females. Nevertheless, they were more successful in raising a brood with higher larval survival compared to pairs that did not experience a food shortage before breeding. We also show that food availability during breeding and social condition had independent effects on the mass of the broods raised, with lighter broods in biparental families than in uniparental ones and on smaller carcasses. Our study thus indicates that a harsh nutritional environment can increase both cooperative as well as competitive interactions between family members. Moreover, our results suggest that it can either hamper or drive the formation of a family because parents choose to restrain reproductive investment in a current brood or are encouraged to breed in a food-poor environment, depending on former experiences and their own nutritional status.     

Alport syndrome caused by a 5′ deletion within the COL4A5 gene

Summary Fourteen Italian patients affected with X-linked Alport syndrome were analyzed by Southern blotting, using cDNA probes of the COL4A5 gene. One proband was shown to carry a large deletion (> 38 kb) that included the 5 part of the gene.     

Repeated Home-Applied Dual-Light Antibacterial Photodynamic Therapy Can Reduce Plaque Burden,Inflammation, and aMMP-8 in Peri-Implant Disease—A Pilot Study

Until now, in clinical dentistry, antibacterial photodynamic therapy (aPDT) has been restricted to in-office treatments, which hampers repeated applications. This pilot study tested the benefit of a commercially available Lumoral^® device designed for regular periodontal dual-light aPDT treatment at home. Seven patients with peri-implant disease applied dual-light aPDT daily in addition to their normal dental hygiene for four weeks. A single Lumoral^® treatment includes an indocyanine green mouth rinse followed by 40 J/cm² radiant exposure to a combination of 810 nm and 405 nm light. A point-of-care analysis of active-matrix metalloproteinase (aMMP-8), visible plaque index (VPI), bleeding on probing (BOP), and peri-implant pocket depth (PPD) measurements was performed on day 0, day 15, and day 30. Reductions in aMMP-8 (p = 0.047), VPI (p = 0.03), and BOP (p = 0.03) were observed, and PPD was measured as being 1 mm lower in the implant (p = ns). These results suggest a benefit of regular application of dual-light aPDT in peri-implantitis. Frequently repeated application can be a promising approach to diminishing the microbial burden and to lowering the tissue destructive proteolytic and inflammatory load around dental implants. Further studies in larger populations are warranted to show the long-term benefits.     

Reproductive success and genetic divergence among varieties of the rare and endangered <Emphasis Type="Italic">Astragalus cremnophylax</Emphasis> (Fabaceae) from Arizona,USA

Populations of Astragalus cremnophylax from Arizona and related A. humillimus from New Mexico were assessed for morphological, reproductive, genetic, and ecological differences. Results suggest that some of the taxa (i.e., A. cremnophylax var. cremnophylax and A. humillimus) are poor reproducers and environmental fluctuation is not likely a factor contributing to low fecundity in these species. Astragalus cremnophylax var. cremnophylax appears to have low fecundity due to inbreeding depression. Allozyme data from 18 loci show little within-population and taxon genetic variability. Most genetic variability is distributed among populations and taxa. The population of A. cremnophylax var. cremnophylax from the Grand Canyon (South Rim) was genetically depauperate, indicative of the bottleneck caused by historic trampling at this viewpoint. Cape Final, a population of A. cremnophylax var. cremnophylax on the Grand Canyon (North Rim) is genetically different from typical materials of this taxon (South Rim) and is more closely related to other taxa within this species complex. However, the other varieties of A. cremnophylax are more closely related to A. humillimus than the population at Cape Final, suggesting potential species status for Cape Final materials based on allozyme divergence.     

Robust parameters confirm predominance of heterotrophic processes in the plankton of a highly humic pond

The conclusions about the role of allochthonous organic matter in the food chains of inland waters are still often conflicting. Here we studied whether the results of specific experiments made in a small pond with very high concentration of allochthonous organic matter could be verified by simple, but robust, basic environmental parameters measured in the field. In summer primary production of phytoplankton could explain only ca. 20% of epilimnetic respiration and <10% of the nutritional requirements of zooplankton. These results agree with those of earlier experimental approaches and unequivocally suggest the major role played by allochthonous nutrition for the plankton in this pond. The roles played by photochemical degradation and anaerobic processes in the transformation of humic compounds available to epilimnetic food web in particular deserve further elucidation.