Microbial biofilms on the sandstone monuments of the Angkor Wat Complex, Cambodia.

Discoloring biofilms from Cambodian temples Angkor Wat, Preah Khan, and the Bayon and West Prasat in Angkor Thom contained a microbial community dominated by coccoid cyanobacteria. Molecular analysis identified Chroococcidiopsis as major colonizer, but low similarity values (<95%) suggested a similar genus or species not present in the databases. In only two of the six sites sampled were filamentous cyanobacteria, Microcoleus, Leptolyngbya, and Scytonema, found; the first two detected by sequencing of 16S rRNA gene library clones from samples of a moist green biofilm on internal walls in Preah Khan, where Lyngbya (possibly synonymous with Microcoleus) was seen by direct microscopy as major colonizer. Scytonema was detected also by microscopy on an internal wall in the Bayon. This suggests that filamentous cyanobacteria are more prevalent in internal (high moisture) areas. Heterotrophic bacteria were found in all samples. DNA sequencing of bands from DGGE gels identified Proteobacteria (Stenotrophomonas maltophilia and Methylobacterium radiotolerans) and Firmicutes (Bacillus sp., Bacillus niacini, Bacillus sporothermodurans, Lysinibacillus fusiformis, Paenibacillus sp., Paenibacillus panacisoli, and Paenibacillus zanthoxyli). Some of these bacteria produce organic acids, potentially degrading stone. Actinobacteria, mainly streptomycetes, were present in most samples; algae and fungi were rare. A dark-pigmented filamentous fungus was detected in internal and external Preah Khan samples, while the alga Trentepohlia was found only in samples taken from external, pink-stained stone at Preah Khan. Results show that these microbial biofilms are mature communities whose major constituents are resistant to dehydration and high levels of irradiation and can be involved in deterioration of sandstone. Such analyses are important prerequisites to the application of control strategies.

Characterization of sulfate-reducing bacteria dominated surface communities during start-up of a down-flow fluidized bed reactor.

An anaerobic down-flow fluidized bed reactor was inoculated with granular sludge and started-up with sulfate containing synthetic wastewater to promote the formation of a biofilm enriched in sulfate-reducing bacteria (SRB), to produce biogenic sulfide. The start-up was done in two stages operating the reactor in batch for 45 days followed by 85 days of continuous operation. Low-density polyethylene was used as support. The biofilm formation was followed up by biochemical and electron microscopy analyses and the composition of the community was examined by 16S rDNA sequence analysis. Maximum immobilized volatile solids (1.2 g IVS/L(support)) were obtained after 14 days in batch regime. During the 85 days of continuous operation, the reactor removed up to 80% of chemical oxygen demand (COD), up to 28% of the supplied sulfate and acetate was present in the effluent. Sulfate-reducing activity determined in the biofilm with ethanol or lactate as substrate was 11.7 and 15.3 g COD/g IVS per day, respectively. These results suggested the immobilization of sulfate reducers that incompletely oxidize the substrate to acetate; the phylogenetic analysis of the cloned 16S rDNA gene sequences showed high identity to the genus Desulfovibrio that oxidizes the substrates incompletely. In contrast, in the granular sludge used as inoculum a considerable number of clones showed homology to Methanobacterium and just few clones were close to SRB. The starting-up approach allowed the enrichment of SRB within the diverse community developed over the polyethylene support.

Biogenic black crusts on buildings in unpolluted environments.

Samples of peeling black crusts from modern and historic buildings in Campeche, Mexico, from a gravestone on the island of Dom Khon, Lao, and from the Anglican cathedral in Belize City were analyzed microbiologically, by scanning electron microscopy plus electron dispersive spectroscopy (EDS) and for pigment composition. In all cases, the surface was covered by a thick mat of cyanobacteria with dark brown sheaths. These were filamentous organisms of the genera Scytonema or Fischerella/Mastigocladus, except for one sample, where coccoid cyanobacteria of Subsection II were predominant. Fungi were not present at all sites and, where seen, were not the major biomass. High scytonemin:chlorophyll a ratios correlated with the dark pigmentation of the cyanobacterial cells and indicated the stressful conditions under which these organisms were living (high temperatures and ultraviolet levels, frequent desiccation). The absence, or low levels, of sulfur in the biofilms confirmed that there was little urban pollution at the sites and the EDS analysis showed that the black coloration was caused solely by cell pigmentation; no dark-colored elements were present at high concentrations. These results demonstrate that, unlike chemically formed thick black crusts found in polluted atmospheres, thin black crusts (which could be called patinas) in clean environments may be predominantly composed of filamentous cyanobacteria.