Comprehensive analysis of different adhesives in aerobiological sampling using optical microscopy and high-throughput DNA sequencing.

The standardization and unification of the procedures to analyze and quantify the airborne pollen concentrations are very important topics. In this work, the effectiveness of the two most used adhesives in aerobiological sampling, silicone prepared with cyclohexane solvent (Silicone) and petroleum jelly (Vaseline), was compared under outdoor conditions. This comparison was carried out using the traditional method based on the identification and quantification by optical microscopy (OM) of the airborne pollen and the novel methodology by high-throughput sequencing analysis (HTS). Globally, the results from both methods of analysis (OM and HTS) showed a good agreement between the two adhesives tested regarding the abundance of the main pollen types present in the samples: Cupressaceae, Olea, Poaceae, Platanus, Quercus. We concluded that the results from both adhesives are comparable data. Furthermore, the comparisons between methodologies, OM vs. HTS, showed that both techniques can accurately identify the most abundant pollen types in the atmosphere for the studied periods, with a good agreement of their relative abundances especially when the airborne pollen diversity is low but showing some divergences as the number of pollen types increases.

Artemisia pollen is the main vector for airborne endotoxin.

BACKGROUND: Endotoxin (LPS) released from gram-negative bacteria causes strong immunologic and inflammatory effects and, when airborne, can contribute to respiratory conditions, such as allergic asthma. OBJECTIVES: We sought to identify the source of airborne endotoxin and the effect of this endotoxin on allergic sensitization. METHODS: We determined LPS levels in outdoor air on a daily basis for 4 consecutive years in Munich (Germany) and Davos (Switzerland). Air was sampled as particulate matter (PM) greater than 10 µm (PM > 10) and PM between 2.5 and 10 µm. LPS levels were determined by using the recombinant Factor C assay. RESULTS: More than 60% of the annual endotoxin exposure was detected in the PM > 10 fraction, showing that bacteria do not aerosolize as independent units or aggregates but adhered to large particles. In Munich 70% of annual exposure was detected between June 12th and August 28th. Multivariate modeling showed that endotoxin levels could be explained by phenological parameters (ie, plant growth). Indeed, days with high airborne endotoxin levels correlated well with the amount of Artemisia pollen in the air. Pollen collected from plants across Europe (100 locations) showed that the highest levels of endotoxin were detected on Artemisia vulgaris (mugwort) pollen, with little on other pollen. Microbiome analysis showed that LPS concentrations on mugwort pollen were related to the presence of Pseudomonas species and Pantoea species communities. In a mouse model of allergic disease, the presence of LPS on mugwort pollen was needed for allergic sensitization. CONCLUSIONS: The majority of airborne endotoxin stems from bacteria dispersed with pollen of only one plant: mugwort. This LPS was essential for inducing inflammation of the lung and allergic sensitization.

Bio-based products control black rot (Xanthomonas campestris pv. campestris) and increase the nutraceutical and antioxidant components in kale.

Black rot of crucifers, (Xanthomonas campestris pv. campestris) is the principal yield-limiting and destructive pathogen of cruciferous crop worldwide. In order to validate a bio-based control alternative for this disease, whey, lime sulfur, biofertilizer, Bordeaux mixture or raw milk were applied to kale (Brassica oleracea var. acephala) plants. The disease control was achieved by most of the tested products. Milk-based products (raw milk and whey) and biofertilizer reduced the severity by 44 and 56% in the field. Antioxidants, crude fibber, crude protein and lipid contents and kale yield were verified in the five treatments on the leaves with and without X. campestris pv. campestris inoculation. In the absence of the pathogen (non-inoculated), lime sulfur and Bordeaux mixture improved plant nutritional value compared to organic treatments, nevertheless milk-based products and biofertilizer improved the evaluated variables more than the control. However, on leaves inoculated with X. campestris pv. campestris raw milk increased antioxidant activity, crude protein and fiber contents, whereas biofertilizer increased kale yield, lipid and antioxidant contents. Milk-based products and biofertilizer were further evaluated in greenhouse trials to determinate the activity of defense-related enzymes and lignin content. Biofertilizer treatment resulted in increased phenylalanine ammonia lyase, catalase, peroxidase activities and lignin content. Hence, the application of milk-based products and biofertilizer are promising to control black rot of crucifers and also improves food quality by boosting nutritional values and antioxidant activity.

Validation of the Hirst-Type Spore Trap for Simultaneous Monitoring of Prokaryotic and Eukaryotic Biodiversities in Urban Air Samples by Next-Generation Sequencing.

Pollen, fungi, and bacteria are the main microscopic biological entities present in outdoor air, causing allergy symptoms and disease transmission and having a significant role in atmosphere dynamics. Despite their relevance, a method for monitoring simultaneously these biological particles in metropolitan environments has not yet been developed. Here, we assessed the use of the Hirst-type spore trap to characterize the global airborne biota by high-throughput DNA sequencing, selecting regions of the 16S rRNA gene and internal transcribed spacer for the taxonomic assignment. We showed that aerobiological communities are well represented by this approach. The operational taxonomic units (OTUs) of two traps working synchronically compiled >87% of the total relative abundance for bacterial diversity collected in each sampler, >89% for fungi, and >97% for pollen. We found a good correspondence between traditional characterization by microscopy and genetic identification, obtaining more-accurate taxonomic assignments and detecting a greater diversity using the latter. We also demonstrated that DNA sequencing accurately detects differences in biodiversity between samples. We concluded that high-throughput DNA sequencing applied to aerobiological samples obtained with Hirst spore traps provides reliable results and can be easily implemented for monitoring prokaryotic and eukaryotic entities present in the air of urban areas.IMPORTANCE Detection, monitoring, and characterization of the wide diversity of biological entities present in the air are difficult tasks that require time and expertise in different disciplines. We have evaluated the use of the Hirst spore trap (an instrument broadly employed in aerobiological studies) to detect and identify these organisms by DNA-based analyses. Our results showed a consistent collection of DNA and a good concordance with traditional methods for identification, suggesting that these devices can be used as a tool for continuous monitoring of the airborne biodiversity, improving taxonomic resolution and characterization together. They are also suitable for acquiring novel DNA amplicon-based information in order to gain a better understanding of the biological particles present in a scarcely known environment such as the air.

Monitoring of airborne biological particles in outdoor atmosphere. Part 1: Importance, variability and ratios.

The first part of this review ("Monitoring of airborne biological particles in outdoor atmosphere. Part 1: Importance, variability and ratios") describes the current knowledge on the major biological particles present in the air regarding their global distribution, concentrations, ratios and influence of meteorological factors in an attempt to provide a framework for monitoring their biodiversity and variability in such a singular environment as the atmosphere. Viruses, bacteria, fungi, pollen and fragments thereof are the most abundant microscopic biological particles in the air outdoors. Some of them can cause allergy and severe diseases in humans, other animals and plants, with the subsequent economic impact. Despite the harsh conditions, they can be found from land and sea surfaces to beyond the troposphere and have been proposed to play a role also in weather conditions and climate change by acting as nucleation particles and inducing water vapour condensation. In regards to their global distribution, marine environments act mostly as a source for bacteria while continents additionally provide fungal and pollen elements. Within terrestrial environments, their abundances and diversity seem to be influenced by the land-use type (rural, urban, coastal) and their particularities. Temporal variability has been observed for all these organisms, mostly triggered by global changes in temperature, relative humidity, et cetera. Local fluctuations in meteorological factors may also result in pronounced changes in the airbiota. Although biological particles can be transported several hundreds of meters from the original source, and even intercontinentally, the time and final distance travelled are strongly influenced by factors such as wind speed and direction. [Int Microbiol 2016; 19(1):1-1 3].