Online Monitoring of Solid-Phase Peptide Syntheses on Glass-Type Surfaces Using White Light Interference

Label-free optical detection methods are of particular value for the investigation of biomolecular interactions. A label-free method based on reflectometric interference spectroscopy is described which enables both the on-line monitoring of solid phase peptide syntheses and subsequently the determination of antibody binding to these peptides without cleavage from the support. The method uses SiO(2) transducers that were modified with diaminopolyethylene glycol. The stepwise coupling of different amino acids to the transducer surface was investigated and the complete monitoring of the synthesis of a viral epitope was performed. The success of the synthesis was proven via binding of a specific monoclonal antibody to the transducer-bound product. In order to demonstrate specificity the binding was inhibited with the same peptide epitope added in solution. The approach is attractive especially in the field of high throughput screening since both the synthesis and the interaction with the biological receptor can be monitored using the same technique.     

Possible Involvement of Altered Gibberellin Metabolism in the Induction of Tomato Irregular Ripening in Dwarf Cherry Tomato by Silverleaf Whitefly

External and internal tomato irregular ripening (TIR) symptoms were associated with the feeding of silverleaf whitefly (SLW), Bemisia argentifolii Bellows and Perring. Four experiments consisting of various soil drench applications of GA₃ (100 ppm) and cycocel (CCC, an inhibitor of GA biosynthesis; 1,000 and 2,000 ppm) were applied to dwarf cherry tomato cv. Florida Petite in the presence and absence of SLW in an attempt to mimic the disorders induced by the SLW. The application of GA₃ induced external and internal TIR symptoms similar to the SLW-induced disorder. Minimal TIR-like symptoms also occurred in the control and CCC treatments. Internal TIR symptoms in GA₃, GA₃ plus SLW, and GA₃ plus CCC treatments ranged from 66% to 97% throughout the experiments. The incidence of external TIR symptoms was highest in the GA₃ plus SLW treatment compared with the other treatments. CCC reduced the incidence of external TIR symptoms induced by GA₃ or GA₃ plus SLW treatments. However, CCC-treated plants also attracted more oviposition and higher populations of SLW and consequently induced a greater incidence of TIR symptoms than SLW treatment alone. Furthermore, although low SLW populations may be associated with low external TIR symptoms, internal TIR symptoms almost always remained high in infested plants. The results suggest that the TIR disorder in dwarf cherry tomato which is induced by the SLW may be a gibberellin-regulated disorder. Received May 27, 1997; accepted September 26, 1997     

Manganese oxidation by microbial consortia from sand filters

The role of microbial consortia on the removal of manganese (Mn) was examined on sand from three different Belgian rapid sand filters for the treatment of ground water. Microorganisms closely associated with deposits of Fe and amorphous Mn precipitates were observed by SEM and EDAX techniques on sand from the filters able to remove Mn efficiently. Bacterial counts were performed. Of the CFU enumerated on PYM-medium, 25–33% displayed Mn-oxidizing activity.Batch cultures were set up by inoculating a Mn-containing, low organic medium with sand from one of the filters. Microbial growth resulted in the formation of Mn-removing bacterial flocs and a pH increase. Suppression of microbial growth by addition of azide, kanamycin, or by autoclaving reduced removal of Mn²⁺ from 0.5 mM/day to 0.05–0.11 mM/day. Buffering the pH of the medium at 7.5 (0.1 mM Hepes) decelerated the Mn removal but did not halt it, whereas microelectrode measurements revealed a clear pH drop of about 0.7 units inside bacterial flocs. In the absence of Mn²⁺, the pH drop was only 0.4 units. The auto-catalytic removal of Mn by the Mn oxide coated filter sand was not sufficient to explain the Mn removal observed. Inactivated cells were not capable of a pronounced autocatalytic Mn removal. Experiments with enrichment cultures indicated that the Mn-removing capacity of the microbial sand filter consortia was not constitutive but was promoted by preadaptation and the presence of a substratum. These results clearly link Mn oxidation in rapid sand filters to microbial processes. Offprint requests to: W. Verstraete.     

Evaluation of some enzymatic methods to measure the bio-activity of aquatic environments

Methods have been evaluated and standardised to measure the phosphatase, saccharase, amylase and lipase enzymatic activities in aquatic environments. The analytical reproducibility, the ecological applicability and the etiology of these activities have been investigated. The usefulness of the various enzymatic assays for routine analyses and for fundamental research of aquatic environments is discussed.     

Selective inhibitors for continuous non-axenic hydrogen production by Rhodobacter capsulatus

To produce H₂ continuously by photosynthetically grown Rhodobacter capsulatus in non-axenic anaerobic reactors, the interaction between the phototroph and possible contaminants was studied and the ecological competitiveness of the Rhodobacter spp. in nitrogen-limited conditions was determined. Experimental test runs showed that blue-green and green algae, sulphate-reducing, acetogenic and methanogenic bacteria significantly interfere with the net amounts of H₂ produced by photobacteria. Therefore, inhibitors to control the growth of those contaminants selectively were screened. By applying a combination of chloroxuron (10mg/l) and cycloheximide (10mg/l) against algae, isohumulones (30 bitterunits/l) and molyb-date (0.5g/l) against sulphate-reducing bacteria and isohumulones and chloroform (10 mg/l) against acetogens and methanogens, photoreactors could be operated in a non-axenic way and continued to produce hydrogen gas at rates depending on the feed quality varying from 333 to 676 ml H₂l reactor/d, for a period of 116d without apparent interference from other microbial contaminants. These findings have a considerable potential for facilitating the isolation of organo-phototrophs and the production of H₂ by these bacteria.     

Mixed culture hydrogenotrophic nitrate reduction in drinking water

Isolation and identification of the bacteria from a hydrogenotrophic reactor for the denitrification of drinking water revealed that several microorganisms are involved. Acinetobacter sp., Aeromonas sp., Pseudomonas sp. and Shewanella putrefaciens were repeatedly isolated from the hydrogenotrophic sludge and postulated to be of primary importance in the process. Nitrate reduction to nitrite appears to be a property of a diverse group of organisms. Nitrite reduction was found to be stimulated by the presence of organic growth factors. Thus, in a mixed culture, hydrogenotrophic denitrification reactor, NO _inf2 ^sup– formed by NO _inf3 ^sup– -reducers can be converted by true denitrifiers thriving on organic growth factors either present in the raw water, or excreted by the microbial community. Mixotrophic growth also contributes to NO _inf2 ^sup– reduction. Finally, chemolithotrophic bacteria participate in the nitrite to nitrogen gas conversion.Offprint requests to: W. Verstraete.     

Competitive ability of amylolytic bacteria in activated sludge.

Shifts were induced into the microbial community of activated sludge by the pulse addition of soluble starch. The subsequent changes of amylolytic and proteolytic microbial populations were recorded. Four amylolytic strains were isolated and characterized with regard to carrying capacity, specific surface and growth kinetics. The competitive ability of these strains was studied by means of two-member competition experiments. These experiments were analysed according to the Lotka-Volterra model and the de Wit method. The different results obtained suggest that the dominance of the amylolytic Pseudomonas sp. (code 01) is based on a combined occurrence of high amylolytic activity, large relative cell surface, high maximum specific growth rate and reduced sensitivity towards associated proteolytic populations.