No abstract

These results demonstrate the efficiency of PMA for differentiating viable and dead P. gingivalis, A. actinomycetemcomitans and F. nucleatum cells, as well as total bacteria, in an in vitro biofilm model, after being exposed to an antimicrobial agent. Hence, this PMA-qPCR method may be useful for studying the effect of antimicrobial agents aimed at oral biofilms.

A bacterial strain capable of growing on propachlor (2-chloro-N-isopropylacetanilide) was isolated from soil by using enrichment and isolation techniques. The strain isolated, designated GCH1, was classified as a member of the genus Pseudomonas. Washed-cell suspensions of strain GCH1 accumulated N-isopropylacetanilide, acetanilide, acetamide, and catechol. Pseudomonas strain GCH1 grew on propachlor with a generation time of 4.2 h and a rate of substrate utilization of 1.75 ؎ 0.15 mol h ؊1 . Gene expression did not require induction but was subject to catabolite expression. Acetanilide was a growth substrate with a yield of 0.56 ؎ 0.02 mg of protein mol ؊1 . GCH1 strain cells were immobilized by adsorption onto a ceramic support and were used as biocatalysts in an immobilized cell system. Propachlor elimination reached 98%, with a retention time of 3 h and an initial organic load of 0.5 mM propachlor. The viability of immobilized cells increased 34-fold after 120 days of bioreactor operation.Propachlor (2-chloro-N-isopropylacetanilide) is an acylanilide herbicide widely used with corn, onion, cabbage, rose bushes, and ornamental plants. Microbial degradation (11,15,16,19) is the primary mechanism of acylanilide dissipation from soil. We previously reported the isolation of Pseudomonas strain PEM1 (3, 10, 11), which metabolizes the herbicide propachlor, yielding N-isopropylacetanilide, acetanilide, and acetamide as intermediates, and the isolation of Acinetobacter strain BEM2, which follows a different pathway and yields N-isopropylaniline and isopropylamine as intermediates. Lee et al. (9) reported that N-isopropylaniline, N-isopropylacetanilide, N-(1-hydroxyisopropyl)acetanilide, and N-isopropyl-2-acetoxyacetanilide were formed in soil treated with propachlor. Villareal et al. (18) proposed a propachlor degradative pathway yielding 2-chloro-N-isopropylacetamide and catechol as intermediates.Groundwater and soil contamination by herbicides has recently become of increasing concern (6,7,20). The relatively new concept of bioremediation provides a potentially cheap alternative to traditional disposal techniques, in addition to representing a genuine removal of contaminants by microbial degradation rather than the relocation of contaminants in such processes as landfilling. The problems presented by continuous fermentation processes could be resolved by the use of immobilized cells as biocatalysts (5,7,12,21).The aim of this study was to characterize the propachlor metabolism of a strain (GCH1) isolated from soil and to test a bioremediation system using immobilized cells in a reactor operating over a period of 150 days. The kinetics of the degradation and the viability of the immobilized cells are reported. MATERIALS AND METHODSIsolation of bacteria. Ten soil samples (10) were collected from agricultural fields in Madrid, Spain, with a history of propachlor contamination. Minimal medium (MB) (4) supplemented with 45 mg of propachlor liter Ϫ1 was inoculated with 20 g of soil sample and incubated at 28°C without sh...

Objectives. The aim of this investigation was to evaluate whether the adenosine triphosphate (ATP) bioluminescence method is an appropriate tool to assess the efficacy of antiseptic mouthrinses in terms of quantitative reductions of total viable microbial counts in mixed biofilm populations in vitro. Study Design. Three mouthrinses, containing respectively, chlorhexidine and cetylpyridinium chloride (CHX/CPC), essential oils (EO) and amine fluoride/stannous fluoride (AFSF), as well as Phosphate Buffered Saline (PBS) used as control, were tested in an in vitro static biofilm model by ATP bioluminescence and compared to culture method. Biofilms were grown on saliva-coated hydroxyapatite disks for 72 hours and then exposed for 1 minute to the mouthrinse or control by immersion. The antibacterial effect of the rinses was tested by analysis of variance. The reliability of the ATP bioluminescence method was assessed by calculating the Pearson correlation coefficients when compared to the viable cell counts obtained by culture. Results. Using ATP bioluminescence, the antimicrobial activity of the tested mouthrinses was demonstrated when compared to the PBS control. The ATP bioluminescence values were significantly correlated (0.769, p<0.001) to the viable cell counts. CHX/CPC and AFSF showed similar antimicrobial activity, although AFSF had a less homogeneous effect, being both more effective than the EO rinse. Conclusion. ATP bioluminescence viability testing may be considered a useful tool to assess the in vitro efficacy of antibacterial compounds. In the proposed model, CHX/CPC and AFSF containing mouthrinses demonstrated superior antimicrobial activity, as compared to EO rinses, in a multispecies biofilm model. Key words:Biofilm, ATP bioluminescence,mouthrinse, essential oils, chlorhexidine, amine fluoride/stannous fluoride.

Klebsiella planticola strain DSZ1 has the ability to degrade different aromatic compounds such as benzoate and organochlorinated as propachlor and alachlor. DSZ1 strain cells mineralised 4-hydroxybenzoate (4HBA) through a meta-cleavage pathway, yielding protocatechuate as dihydroxylated intermediate, with a specific rate of CO2 formation 0.12 x 10-6 (cpm/OD) h-1, and a rate of 4-HBA utilisation of 0.75 mmol h-1. Aerobically the 4HBA transport system is driven by gradient of protons (DeltapH), but is not ATP-driven. Under anaerobic conditions, the system can use the nitrate reduction as a final electron acceptor in respiration. A kinetic analysis of the 4HBA transport system revealed a Kt value of 16 microM with a Vmax value of 25 nmol/min.mg at pH 7.