Martha Hengst

Bacterial Active Community Cycling in Response to Solar Radiation and Their Influence on Nutrient Changes in a High-Altitude Wetland

Microbial communities inhabiting high-altitude spring ecosystems are subjected to extreme changes in solar irradiance and temperature throughout the diel cycle. Here, using 16S rRNA gene tag pyrosequencing (cDNA) we determined the composition of actively transcribing bacteria from spring waters experimentally exposed through the day (morning, noon, and afternoon) to variable levels of solar radiation and light quality, and evaluated their influence on nutrient recycling. Solar irradiance, temperature, and changes in nutrient dynamics were associated with changes in the active bacterial community structure, predominantly by Cyanobacteria, Verrucomicrobia, Proteobacteria, and 35 other Phyla, including the recently described Candidate Phyla Radiation (e.g., Parcubacteria, Gracilibacteria, OP3, TM6, SR1). Diversity increased at noon, when the highest irradiances were measured (3.3–3.9 H′, 1125 W m-2) compared to morning and afternoon (0.6–2.8 H′). This shift was associated with a decrease in the contribution to pyrolibraries by Cyanobacteria and an increase of Proteobacteria and other initially low frequently and rare bacteria phyla (< 0.5%) in the pyrolibraries. A potential increase in the activity of Cyanobacteria and other phototrophic groups, e.g., Rhodobacterales, was observed and associated with UVR, suggesting the presence of photo-activated repair mechanisms to resist high levels of solar radiation. In addition, the percentage contribution of cyanobacterial sequences in the afternoon was similar to those recorded in the morning. The shifts in the contribution by Cyanobacteria also influenced the rate of change in nitrate, nitrite, and phosphate, highlighted by a high level of nitrate accumulation during hours of high radiation and temperature associated with nitrifying bacteria activity. We did not detect ammonia or nitrite oxidizing bacteria in situ, but both functional groups (Nitrosomona and Nitrospira) appeared mainly in pyrolibraries generated from dark incubations. In total, our results reveal that both the structure and the diversity of the active bacteria community was extremely dynamic through the day, and showed marked shifts in composition that influenced nutrient recycling, highlighting how abiotic variation affects potential ecosystem functioning.

Microbial activity response to solar radiation across contrasting environmental conditions in Salar de Huasco, northern Chilean altiplano

In high altitude environments, extreme levels of solar radiation and important differences of ionic concentrations over narrow spatial scales may modulate microbial activity. In Salar de Huasco, a high-altitude wetland in the Andean mountains, the high diversity of microbial communities has been characterized and associated with strong environmental variability. Communities that differed in light history and environmental conditions, such as nutrient concentrations and salinity from different spatial locations, were assessed for bacterial secondary production (BSP, 3H-leucine incorporation) response from short-term exposures to solar radiation. We sampled during austral spring seven stations categorized as: (a) source stations, with recently emerged groundwater (no-previous solar exposure); (b) stream running water stations; (c) stations connected to source waters but far downstream from source points; and (d) isolated ponds disconnected from ground sources or streams with a longer isolation and solar exposure history. Very high values of 0.25 μE m-2 s-1, 72 W m-2 and 12 W m-2 were measured for PAR, UVA, and UVB incident solar radiation, respectively. The environmental factors measured formed two groups of stations reflected by principal component analyses (near to groundwater sources and isolated systems) where isolated ponds had the highest BSP and microbial abundance (35 microalgae taxa, picoeukaryotes, nanoflagellates, and bacteria) plus higher salinities and PO43- concentrations. BSP short-term response (4 h) to solar radiation was measured by 3H-leucine incorporation under four different solar conditions: full sun, no UVB, PAR, and dark. Microbial communities established in waters with the longest surface exposure (e.g., isolated ponds) had the lowest BSP response to solar radiation treatments, and thus were likely best adapted to solar radiation exposure contrary to ground source waters. These results support our light history (solar exposure) hypothesis where the more isolated the community is from ground water sources, the better adapted it is to solar radiation. We suggest that factors other than solar radiation (e.g., salinity, PO43-, NO3-) are also important in determining microbial productivity in heterogeneous environments such as the Salar de Huasco.

Bacterial Survival under Extreme UV Radiation: A Comparative Proteomics Study of Rhodobacter sp., Isolated from High Altitude Wetlands in Chile

Salar de Huasco, defined as a polyextreme environment, is a high altitude saline wetland in the Chilean Altiplano (3800 m.a.s.l.), permanently exposed to the highest solar radiation doses registered in the world. We present here the first comparative proteomics study of a photoheterotrophic bacterium, Rhodobacter sp., isolated from this remote and hostile habitat. We developed an innovative experimental approach using different sources of radiation (in situ sunlight and UVB lamps), cut-off filters (Mylar, Lee filters) and a high-throughput, label-free quantitative proteomics method to comprehensively analyze the effect of seven spectral bands on protein regulation. A hierarchical cluster analysis of 40 common proteins revealed that all conditions containing the most damaging UVB radiation induced similar pattern of protein regulation compared with UVA and visible light spectral bands. Moreover, it appeared that the cellular adaptation of Rhodobacter sp. to osmotic stress encountered in the hypersaline environment from which it was originally isolated, might further a higher resistance to damaging UV radiation. Indeed, proteins involved in the synthesis and transport of key osmoprotectants, such as glycine betaine and inositol, were found in very high abundance under UV radiation compared to the dark control, suggesting the function of osmolytes as efficient reactive oxygen scavengers. Our study also revealed a RecA-independent response and a tightly regulated network of protein quality control involving proteases and chaperones to selectively degrade misfolded and/or damaged proteins.

Microbial diversity and trophic components of two high altitude wetlands of the Chilean Altiplano

Este estudio examina la limnologia y ecologia de dos lagos de altura, los humedales de Lirima (19°51’24 S; 68°55’02 W; 4000 m asl) y Caya (20°37’21 S; 68°58’28 W; 3700 m asl) que estan ubicados en el Altiplano Chileno, representando cuencas evaporiticas remanentes de antiguos paleolagos los cuales ocuparon una amplia zona en lo que hoy conocemos como Altiplano. Estos sistemas tienen un balance hidrico negativo, recibiendo su agua desde fuentes freaticas, derretimiento de nueves y escasas lluvias estacionales. Las caracteristicas del suelo junto con la demanda de agua en la region han acelerado el proceso desalinizacion en estos sistemas siendo reflejado en las caracteristicas quimicas actuales. Los valores de nutrientes fueron tipicos de sistemas mesotroficos a eutroficos. El contenido ionico clasifica Lirima como un humedal sodio sulfatado y el humedal de Caya como un humedal de cloruro de calcio. Los valores de conductividad fluctuaron entre 778 μS/cm en Lirima a 2100 μS/cm en Caya, reflejandose en las diferencias de biodiversidad encontrada en estos sistemas. El humedal de Lirima conserva el pez endemico Orestias que representa Unidades Evolutivas Significativas (ESU) en la region. La diversidad microbiana en muestras de agua estuvo caracterizada por la presencia de 5 filo bacterianos y generos relacionados (e.g. Psychrobacter, Bacillus, Eryhtobacter, Halomonas). Los componentes troficos estudiados incluyeron macrofitas, plancton, bentos, peces, anfibios y aves. Este articulo descriptivo destaca las inusuales caracteristicas limnologicas y biologicas de los humedales de altura poniendo atencion a la importancia de describir comunidades en distintos niveles de organizacion biologica (desde tapetes microbianos hasta vertebrados superiores), pero tambien sus funciones, interacciones y sensibilidad a cambios en la disponibilidad de agua.

Active microbiome structure and its association with environmental factors and viruses at different aquatic sites of a high-altitude wetland

Salar de Huasco is a high‐altitude wetland characterized by a highly diverse microbial life adapted to extreme climatic and environmental conditions. Our study aims to determine active microbial community structure changes within different aquatic sites and its relationship with environmental factors and viruses as potential drivers of diversification in different aquatic areas of this ecosystem. In this study, bacteria and archaea composition (16S rRNA subunit pyrolibraries) and picoplankton and viral abundance were determined at ponds, springs and lagoon sites of the wetland during wet and dry seasons (February and July 2012, respectively). In general, mixosaline waters (1,400–51,000 μS/cm) usually found in ponds and lagoon presented higher picoplanktonic abundances compared to freshwater (<800 μS/cm) spring sites, ranging from 1.07 × 105 to 1.83 × 107 cells/ml. Viral abundance and viral to picoplankton ratio (VPR) also presented greater values at ponds compared to spring sites, reaching up to 4.78 × 108 viruses‐like particles and up to 351 for VPR. In general, ponds hold a higher microbial diversity and complexity associated also with the presence of microbial mats compared with water sources or lagoon (Shannon index H′ 2.6–3.9 vs. <2.0). A greater richness of archaea was also detected in ponds characterized by functional groups such as known methanogens and ammonia oxidizers, and uncultured groups. In total, our results indicate that among the different aquatic sites of the wetland, ponds presented a great microbial community diversification associated to a higher top‐down control by viruses which may influence nutrient and greenhouse gases cycling.

The activity of nitrifying microorganisms in a high-altitude Andean wetland

High-altitude wetland holds freshwater springs, evaporitic ponds and lagoon with variable salinity and nutrients, potentially influencing the ecology of nitrifying communities. In this study, nitrifying microorganisms in Salar de Huasco (Chile) were surveyed to determine bacterial and archaeal contribution to ammonium (AO), nitrite oxidation (NO), ammonium uptake (AU) during wet and dry seasons. The activity signals from these groups were assessed by specific amoA-qPCR transcription, 15N tracer studies and addition of group specific inhibitor experiments for nitrifying microorganisms (N1-guanyl-1, 7-diaminoheptane [GC7]-archaeal specific and allylthiourea [ATU]-bacterial specific). Nitrifying communities, i.e. Nitrosopumilus, Nitrosospira, Nitrosomonas, Kuenenia and Nitrospira, were more frequent (∼0.25% of 16S rRNA sequences) at low salinity sites. Bacterial amoA-qPCR transcripts also increased at low salinity and along in situ ammonium increase observed between wet/dry seasons. Nutrient changes through time and 15N tracer experiments results showed that AO and NO were detected and peaked mainly at low salinity-high ammonium sites (<37 000 μS cm-1 and >0.3 μM), whereas AU was predominant at evaporitic sites. Our results indicate that salinity and ammonium affect the nitrifying communities that are potentially more active at low-salinity sites but persistent at saltier evaporitic areas of the wetland when ammonium is available.

Microbial community composition and trophic role along a marked salinity gradient in Laguna Puilar, Salar de Atacama, Chile

The geological, hydrological and microbiological features of the Salar de Atacama, the most extensive evaporitic sedimentary basin in the Atacama Desert of northern Chile, have been extensively studied. In contrast, relatively little attention has been paid to the composition and roles of microbial communities in hypersaline lakes which are a unique feature in the Salar. In the present study biochemical, chemical and molecular biological tools were used to determine the composition and roles of microbial communities in water, microbial mats and sediments along a marked salinity gradient in Laguna Puilar which is located in the “Los Flamencos” National Reserve. The bacterial communities at the sampling sites were dominated by members of the phyla Bacteroidetes, Chloroflexi, Cyanobacteria and Proteobacteria. Stable isotope and fatty acid analyses revealed marked variability in the composition of microbial mats at different sampling sites both horizontally (at different sites) and vertically (in the different layers). The Laguna Puilar was shown to be a microbially dominated ecosystem in which more than 60% of the fatty acids at particular sites are of bacterial origin. Our pioneering studies also suggest that the energy budgets of avian consumers (three flamingo species) and dominant invertebrates (amphipods and gastropods) use minerals as a source of energy and nutrients. Overall, the results of this study support the view that the Salar de Atacama is a heterogeneous and fragile ecosystem where small changes in environmental conditions may alter the balance of microbial communities with possible consequences at different trophic levels.

Diversity of Thermophilic Iron-Pyrite-Oxidizing Enrichments from Solfataric Hot Springs in the Chilean Altiplano

Relatively little is known about the microbial communities present in natural environments that meet physico-chemical conditions for the development of potential leaching microorganisms such as thermal ecosystems from the Chilean Altiplano. Thermophilic leaching enrichments were obtained and identified from a high altitude solfataric pound in Lirima hot springs in the Chilean Altiplano. This ecosystem is characterized by hot underground freshwaters, enriched in sulfur compounds showing pH from neutral to acidic. Microbial diversity has been scarcely explored here, and preliminary results demonstrate that hydrothermal pounds are represented by thermophilic anaerobic and acidophilic taxa. Thermophilic leaching cultures in shake flasks were obtained using ferrous iron and pyrite as energy source. The presence of Bacteria and Archaea in oxidizing enrichments was determined by PCR amplification of 16S rRNA genes. A preliminary analysis of microbial diversity using massive sequencing revealed that Bacteria were more abundant than Archaea in both enrichments. Specifically for the iron-oxidizing culture, the majority of the sequences clustered within the Proteobacteria phylum (79%). Among Proteobacteria, the proportion of Betaproteobacteria (42.2%) and Gammaproteobacteria (21.27%) was much higher than that of Alphaproteobacteria (15.5%). Within the Betaproteobacteria class, the most frequent genus was Leptothrix-like. Similar results were obtained for the pyrite oxidizing culture. Interestingly, this study shows the presence of microorganisms close to the Leptothrix genus under low pH conditions (1.7-2.8) and their capacity to grow at high temperatures with ferrous iron or pyrite as sole energy source

A Functional Perspective Analysis of Macroalgae and Epiphytic Bacterial Community Interaction

Macroalgae are photosynthetic, multicellular, sessile eukaryotic organisms that offer diverse habitats for the colonization of epiphytic bacteria, therefore establishing biological interactions of diverse complexity. This review focusses on the interactions between macroalgae and their Epiphytic Bacterial Community (EBC); the main aims are to ascertain whether (1) the epiphytic bacterial groups differ at the phylum and genus levels of the macroalgae; (2) the methodologies used so far to study these microorganisms are related in any way to eventual variations of the EBCs on macroalgae; and (3) the EBC of macroalgae has a functional means rather a simple taxonomic grouping. Results showed firstly the taxonomic grouping of macroalgae does not explain the composition and structure of the EBCs. Secondly, the methodology used is important for describing EBCs; and thirdly, multiple bacteria can have the same function and thus to describe the functionality of EBCs it is important to recognize host-specific and generalist bacteria. We recommend the incorporation of a complementary approach between the taxonomic composition and the functional composition analyzes of EBCs, as well as the use of methodological tools that allow analysis of interactions between the EBCs and their hosts, based on the “holobiont” concept.

Revisión del género Philonerax Bromley, 1932 (Diptera: Asilidae: Asilinae) y dos nuevas especies

El genero Philonerax descrito por Bromley en 1932, fue sinonimizado a Lochmorhynchus Engel, 1930 por Artigas & Papavero (1995). En este trabajo, Philonerax recupera su estatus de genero; la especie Philonerax cribatus Hull es sinonimizada a Ph. mucidus (Walker) y se crean dos nueva especies: Ph. amblayoensis sp. nov. de Argentina y Ph. cienagaensis sp. nov. de Argentina y Bolivia; principalmente en base a caracteres de la genitalia de machos y hembras