A. Fernandez-Vivas

Bioconservation of deteriorated monumental calcarenite stone and identification of bacteria with carbonatogenic activity

The deterioration of the stone built and sculptural heritage has prompted the search and development of novel consolidation/protection treatments that can overcome the limitations of traditional ones. Attention has been drawn to bioconservation, particularly bacterial carbonatogenesis (i.e. bacterially induced calcium carbonate precipitation), as a new environmentally friendly effective conservation strategy, especially suitable for carbonate stones. Here, we study the effects of an in situ bacterial bioconsolidation treatment applied on porous limestone (calcarenite) in the sixteenth century San Jeronimo Monastery in Granada, Spain. The treatment consisted in the application of a nutritional solution (with and without Myxococcus xanthus inoculation) on decayed calcarenite stone blocks. The treatment promoted the development of heterotrophic bacteria able to induce carbonatogenesis. Both the consolidation effect of the treatment and the response of the culturable bacterial community present in the decayed stone were evaluated. A significant surface strengthening (consolidation) of the stone, without altering its surface appearance or inducing any detrimental side effect, was achieved upon application of the nutritional solution. The treatment efficacy was independent of the presence of M. xanthus (which is known as an effective carbonatogenic bacterium). The genetic diversity of 116 bacterial strains isolated from the stone, of which 113 strains showed carbonatogenic activity, was analysed by repetitive extragenic palindromic–polymerase chain reaction (REP-PCR) and 16S rRNA gene sequencing. The strains were distributed into 31 groups on the basis of their REP-PCR patterns, and a representative strain of each group was subjected to 16S rRNA gene sequencing. Analysis of these sequences showed that isolates belong to a wide variety of phylogenetic groups being closely related to species of 15 genera within the Proteobacteria, Firmicutes and the Actinobacteria. This study shows that the abundant carbonatogenic bacteria present in the decayed stone are able to effectively consolidate the degraded stone by producing new calcite (and vaterite) cement if an adequate nutritional solution is used. The implications of these results for the conservation of cultural heritage are discussed.

Biotransformation of oleanolic and maslinic acids by Rhizomucor miehei.

Microbial transformation of oleanolic acid by Rhizomucor miehei produced three metabolites. A known compound, a 30-hydroxyl derivative (queretaroic acid), and two 7β,30- and 1β,30-dihydroxylated metabolites, respectively. The action of the same fungus (R. miehei) on maslinic acid produced an olean-11-en-28,13β-olide derivative, a metabolite hydroxylated at C-30, an 11-oxo derivative, and two metabolites with an 11α,12α-epoxy group, hydroxylated or not at C-30. Their structures were elucidated by extensive analyses of their spectroscopic data, and also by chemical correlations.

Evidence for an activating substance related to autolysis in Myxococcus coralloides D

When cells of Myxococcus coralloides D grow as a suspension in liquid media the cultures enter a lysis phase after having reached a critical level of cell density. Supernatants of prelytic cells added to fresh cultures are able to induce autolysis quickly. If prelytic cells are transferred to supernatants of fresh cultures, autolysis is retarded; thus delay is also observed when prelytic cells are transferred to fresh medium at the same cellular density. In the sequence of events resulting in cell lysis an activating substance may be involved. This substance may be excreted into the medium. The substance seems to be stable at high temperatures, extreme pH values and enzymatic treatment.

Biotransformation of oleanolic and maslinic methyl esters by Rhizomucor miehei CECT 2749

The pentacyclic triterpenoids methyl oleanolate, methyl maslinate, methyl 3β-hydroxyolean-9(11),12-dien-28-oate, and methyl 2α,3β-dihydroxy-12β,13β-epoxyolean-28-oate were biotransformed by Rhizomucor miehei CECT 2749. Microbial transformation of methyl oleanolate produced only a 7β,30-dihydroxylated metabolite with a conjugated 9(11),12-diene system in the C ring. Biotransformation of the substrate with this 9(11),12-diene system gave the same 7β,30-dihydroxylated compound together with a 7β,15α,30-trihydroxyl derivative. The action of this fungus (R. miehei) on methyl maslinate was more varied, isolating metabolites with a 30-hydroxyl group, a 9(11),12-diene system, an 11-oxo group, or an 12-oxo group. Microbial transformation of the substrate with a 12β,13β-epoxy function resulted in the isolation of two metabolites with 12-oxo and 28,13β-olide groups, hydroxylated or not at C-7β, together with a 30-hydroxy-12-oxo derivative. The structures of these derivatives were deduced by extensive and rigorous spectroscopic studies.

Polyphosphate glucokinase and ATP glucokinase activities in Mycococcus coralloides D

Polyphosphate glucokinase and ATP glucokinase were detected in cell-free extracts of Myxococcus coralloides strain D, but pyrophosphate glucokinase was not detected. Both glucokinase activities were separated by chromatography. The approximate molecular weight is 61 000 for polyphosphate glucokinase and 47 000 for ATP glucokinase. Substrate specificity and pH optimum was studied in the polyphosphate glucokinase. Polyphosphate and ATP glucokinase activities were verified by 13C Nuclear Magnetic Resonance.