Selection of LED lighting systems for the reduction of the biodeterioration of speleothems induced by photosynthetic biofilms in the Nerja Cave (Malaga, Spain).

Electrical lighting favours the development of photosynthetic biofilms in caves which can induce biodeterioration in the colonized substrates. The use of specific lights as a limiting factor for biofilm growth could be effective in their control and represents an alternative to chemical methods since they can damage the substrate. However, studies about lighting and the photosynthetic activity of organisms in caves are scarce. In order to select the most effective LED light source in reducing photosynthesis and therefore, in reducing the growth rates of microalgae and cyanobacteria, four biofilms in the Nerja Cave were illuminated by several light emitted diodes (LEDs) with different spectral compositions and the photobiological responses were measured both by empirical and theoretical methodologies. The empirical approach was based on the photosynthetic efficiency, by measuring the in vivo chlorophyll a (Chl a) fluorescence and the theoretical approach was based on the photonic assimilation performance related to the proportion of the light quality used for photosynthesis, according to the action spectra for photosynthesis available in the literature. The photobiological responses showed differences between the empirical and theoretical approach mainly in biofilms dominated by cyanobacteria and red algae, probably because the available action spectra were not useful for monitoring these Nerja Cave biofilms. However, the expected spectral responses of photosynthesis were observed in green microalgal biofilms with maximum photosynthetic efficiency in red and blue light although the green light was also unexpectedly high. The high photosynthetic efficiency in green light could be explained by the predictable high chlorophyll content due to a very dark environment. The results were not conclusive enough for all the biofilm types to be able to recommend a specific lighting system for the photocontrol of biofilm expansion. Therefore, new action spectra for photosynthesis of the extremophile organisms of the Nerja Cave are required. This approach, based on theoretical and empirical methodologies, is a useful tool to obtain information to allow the design of the most adequate lighting systems to reduce photosynthetic activity and favour the conservation of the caves.

Chalicogloea cavernicola gen. nov., sp. nov. (Chroococcales, Cyanobacteria), from low-light aerophytic environments: combined molecular, phenotypic and ecological criteria.

This work characterizes a unicellular cyanobacterium with nearly spherical cells and thin-outlined sheaths that divide irregularly, forming small packets immersed in a diffluent mucilaginous layer. It was isolated growing on calcite speleothems and walls in a show cave in Collbató (Barcelona, Spain). Spectral confocal laser and transmission electron microscopy were used to describe the morphology, fine structure and thylakoid arrangement. The pigments identified were phycoerythrin, phycocyanin, allophycocyanin and chlorophyll a. Three-dimensional reconstructions, generated from natural fluorescence z-stacks, revealed a large surface area of nearly flat, arm-like thylakoidal membranes connected to each other and forming a unified structure in a way that, to our knowledge, has never been described before. Phylogenetic analyses using the 16S rRNA gene sequence showed 95% similarity to strain Chroococcus sp. JJCM (GenBank accession no. AM710384). The diacritical phenotypic features do not correspond to any species currently described, and the genetic traits support the strain being classified as the first member of an independent genus in the order Chroococcales and the family Chroococcaceae. Hence, we propose the name Chalicogloea cavernicola gen. nov., sp. nov. under the provisions of the International Code of Nomenclature for Algae, Fungi and Plants. The type strain of Chalicogloea cavernicola is COLL 3(T) (=CCALA 975(T) =CCAP 1424/1(T)).

Enigmatic reticulated filaments in subsurface granite.

In the last few years, geomicrobiologists have focused their researches on the nature and origin of enigmatic reticulated filaments reported in modern and fossil samples from limestone caves and basalt lava tubes. Researchers have posed questions on these filaments concerning their nature, origin, chemistry, morphology, mode of formation and growth. A tentative microbial origin has been elusive since these filaments are found as hollow tubular sheaths and could not be affiliated to any known microorganism. We describe the presence of similar structures in a 16th century granite tunnel in Porto, Northwest Portugal. The reticulated filaments we identify exhibit fine geometry surface ornamentation formed by cross-linked Mn-rich nanofibres, surrounded by a large amount of extracellular polymeric substances. Within these Mn-rich filaments we report for the first time the occurrence of microbial cells.

Morphology and molecular evaluation of Iphinoe spelaeobios gen. nov., sp. nov. and Loriellopsis cavernicola gen. nov., sp. nov., two stigonematalean cyanobacteria from Greek and Spanish caves.

Caves have generally been found to host phototrophic micro-organisms from various taxonomic groups, with cyanobacteria comprising an important group that have adapted to these stable and highly specific environments. A polyphasic study based on aspects of classical morphology and molecular data revealed two new monospecific genera from fresh material of Greek and Spanish caves. Both taxa are characterized by obligatory true branching (T-type, V-type and false branching), the presence of heterocysts, and reproduction by hormocysts and akinetes. They shared some similarities in their morphological characteristics as revealed by light, scanning electron and transmission electron microscopy, but phylogenetic analysis based on 16S rRNA gene sequences showed that the two phylotypes were different (89.8% similarity); this represents an example of shared morphology in genetically different strains of cave-adapted species. Phenotypic and genetic traits strongly support classification of the phylotypes as independent taxa in the order Stigonematales (the most differentiated and complicated group of cyanobacteria), family Loriellaceae Geitl 1925. Hence, the names Iphinoe spelaeobios Lamprinou and Pantazidou gen. nov., sp. nov. and Loriellopsis cavernicola Hernández-Mariné and Canals gen. nov., sp. nov. are proposed.

Laboratory-induced endolithic growth in calcarenites: biodeteriorating potential assessment.

This study is aimed to assess the formation of photosynthetic biofilms on and within different natural stone materials, and to analyse their biogeophysical and biogeochemical deterioration potential. This was performed by means of artificial colonisation under laboratory conditions during 3 months. Monitoring of microbial development was performed by image analysis and biofilm biomass estimation by chlorophyll extraction technique. Microscopy investigations were carried out to study relationships between microorganisms and the mineral substrata. The model applied in this work corroborated a successful survival strategy inside endolithic microhabitat, using natural phototrophic biofilm cultivation, composed by cyanobacteria and algae, which increased intrinsic porosity by active mineral dissolution. We observed the presence of mineral-like iron derivatives (e.g. maghemite) around the cells and intracellularly and the precipitation of hausmannite, suggesting manganese transformations related to the biomineralisation.

Cyanobacteria-containing biofilms from a Mayan monument in Palenque, Mexico.

Surfaces of buildings at the archaeological site of Palenque, Mexico, are colonized by cyanobacteria that form biofilms, which in turn cause aesthetic and structural damage. The structural characterization and species composition of biofilms from the walls of one of these buildings, El Palacio, are reported. The distribution of photosynthetic microorganisms in the biofilms, their relationship with the colonized substratum, and the three-dimensional structure of the biofilms were studied by image analysis. The differences between local seasonal microenvironments at the Palenque site, the bioreceptivity of stone and the relationship between biofilms and their substrata are described. The implications for the development and permanence of species capable of withstanding temporal heterogeneity in and on El Palacio, mainly due to alternating wet and dry seasons, are discussed. Knowledge on how different biofilms contribute to biodegradation or bioprotection of the substratum can be used to develop maintenance and conservation protocols for cultural heritage.

Does green light influence the fluorescence properties and structure of phototrophic biofilms?

Artificial illumination can harm works of art by inducing the development of photosynthetic biofilms. With the aim of preventing biodeterioration or esthetic damage to such surfaces, we evaluated and compared the effects of illuminating biofilms formed by Gloeothece membranacea (cyanobacteria) and Chlorella sorokiniana (Chlorophyta) using exclusively white or green light.

Long term diurnal variations in contaminant removal in high rate ponds treating urban wastewater.

In this investigation, diurnal variations in contaminant removal in high rate ponds (HRP) treating urban wastewater were evaluated. Two experimental HRPs (surface area 1.54 m2 and depth 0.3 m), each with a clarifier in series (surface area 0.025 m2), were operated in parallel with different hydraulic retention times (3-10 days) but with the same environmental conditions over a period of one year. The operating strategies adopted only yielded a significant overall difference in removal between the two HRPs for nutrients. Effluent total suspended solids and chemical oxygen demand were slightly higher at midday than at dawn, while for total nitrogen and total phosphorous the concentrations were slightly higher at dawn. All these differences were related to the diurnal changes of DO and pH. The main conclusion of this work is that the diurnal variations of the contaminant concentrations in HRPs do not seriously affect their reliability in treating wastewater.

Phyllobacterium catacumbae sp. nov., a member of the order 'Rhizobiales' isolated from Roman catacombs.

Two strains were isolated from tuff, a volcanic rock that forms the walls of the Roman Catacombs of Saint Callixtus in Rome, Italy. A polyphasic approach using nutritional and physiological tests, reactions to antibiotics, fatty acid profiles, DNA base ratios, DNA-DNA reassociation and 16S rRNA gene sequence comparisons showed that the two isolates belong to a novel species within the genus Phyllobacterium. The species Phyllobacterium catacumbae sp. nov. is proposed. The type strain is CSC19(T) (=CECT 5680(T)=LMG 22520(T)).

Noninvasive pigment identification in single cells from living phototrophic biofilms by confocal imaging spectrofluorometry.

A new imaging technique for the analysis of fluorescent pigments from a single cell is reported. It is based on confocal scanning laser microscopy coupled with spectrofluorometric methods. The setup allows simultaneous establishment of the relationships among pigment analysis in vivo, morphology, and three-dimensional localization inside thick intact microbial assemblages.

Unusual ultrastructural features in three strains of Cyanothece (cyanobacteria).

Three unicellular cyanobacterial strains (PCC 7425, PCC 8303, PCC 9308) assigned to the genus Cyanothece Komárek 1976, which showed an unusually high content of light refractile inclusions when viewed by phase-contrast microscopy, were characterized by confocal laser scanning microscopy and transmission electron microscopy. All strains had concentric cortical thylakoids and a compact central nucleoid. Frequently, the two innermost thylakoid membranes protruded to form circular enclosures containing cytoplasm or electron-transparent granules, or both. The largest granules were partially immersed in the nucleoid region, but they remained attached to the inner cortical thylakoids by a single narrow connection. The pattern of binary cell division in strain PCC 7425 was different than that in strains PCC 8303 and PCC 9308. In the former, all cell wall layers invaginated simultaneously, whereas in the latter the invagination of the outer membrane was delayed compared to that of the cytoplasmic membrane and the peptidoglycan layer. Thus, prior to completion of cell division, the new daughter cells of strains PCC 8303 and PCC 9308 were transiently connected by a thick septum, which was not observed in strain PCC 7425. Nucleoid partitioning coincided with initiation of cell division in all three strains and was unlike that reported in other bacteria and in archaea, in which separation of the nucleoids precedes cell division. Based on the common morphological and ultrastructural features, the three strains of Cyanothece examined constitute a distinct cluster, which might deserve independent generic status.