Ice Nucleation Activity Research Articles

Phytoliths in particulate matter released by wind erosion on arable land in La Pampa, Argentina

Silicon (Si) is considered a beneficial element in plant nutrition, but its importance on ecosystems goes far beyond that. Various forms of silicon are found in soils, of which the phytogenic pool plays a decisive role due to its good availability. This Si returns to the soil through the decomposition of plant residues, where they then participate in the further cycle as biogenic amorphous silica (bASi) or so-called phytoliths. These have a high affinity for water, so that the water holding capacity and water availability of soils can be increased even by small amounts of ASi. Agricultural land is a considerable global dust source, and dust samples from arable land have shown in cloud formation experiments a several times higher ice nucleation activity than pure mineral dust. Here, particle sizes in the particulate matter fractions (PM) are important, which can travel long distances and reach high altitudes in the atmosphere. Based on this, the research question was whether phytoliths could be detected in PM samples from wind erosion events, what are the main particle sizes of phytoliths and whether an initial quantification was possible.Measurements of PM concentrations were carried out at a wind erosion measuring field in the province La Pampa, Argentina. PM were sampled during five erosion events with Environmental Dust Monitors (EDM). After counting and classifying all particles with diameters between 0.3 and 32 µm in the EDMs, they are collected on filters. The filters were analyzed by Scanning Electron Microscopy and Energy Dispersive X-Ray analysis (SEM-EDX) to investigate single or ensembles of particles regarding composition and possible origins.The analyses showed up to 8.3 per cent being phytoliths in the emitted dust and up to 25 per cent of organic origin. Particles of organic origin are mostly in the coarse dust fraction, whereas phytoliths are predominately transported in the finer dust fractions. Since phytoliths are both an important source of Si as a plant nutrient and are also involved in soil C fixation, their losses from arable land via dust emissions should be considered and its specific influence on atmospheric processes should be studied in detail in the future.

Ice-nucleating particles near two major dust source regions

Abstract. Mineral dust and sea spray aerosol represent important sources of ice-nucleating particles (INPs), the minor fraction of aerosol particles able to trigger cloud ice crystal formation and, consequently, influence multiple climate-relevant cloud properties including lifetime, radiative properties and precipitation initiation efficiency. Mineral dust is considered the dominant INP source in many parts of the world due to its ice nucleation efficiency and its sheer abundance, with global emission rates of up to 4700 Tg a−1. However, INPs emitted from the ocean surface in sea spray aerosol frequently dominate INP populations in remote marine environments, including parts of the Southern Ocean where cloud-resolving model simulations have demonstrated that cloud radiative properties are likely strongly controlled by INPs. Here we report INP concentrations measured in aerosol and seawater samples during Air Quality and Climate Change in the Arabian Basin (AQABA), a shipborne campaign that spanned the Red Sea, Gulf of Aden, Arabian Sea, Arabian Gulf and part of the Mediterranean. In aerosol samples collected within a few hundred kilometers of the first and second ranked sources of dust globally, the Sahara and Arabian Peninsula, INP concentrations ranged from 0.2 to 11 L−1 at −20 ∘C with observed ice-active surface site densities (ns) 1–3 orders of magnitude below levels predicted by mineral dust INP parameterizations. Over half of the samples (at least 14 of 26) were collected during dust storms with average dust mass concentrations between 150 and 490 µg m−3 (PM10), as simulated by the Modern-Era Retrospective analysis for Research and Application, version 2 (MERRA-2). The impacts of heat and peroxide treatments indicate that organics dominated the observed ice nucleation (IN) activity at temperatures ≥ −15 ∘C with proteinaceous (heat-labile) INPs frequently observed at high freezing temperatures >−10 ∘C. INP concentrations in seawater samples ranged between 3 and 46 mL−1 at −19 ∘C, demonstrating the relatively low INP source potential of seawater in the region as compared to seawater from multiple other regions reported previously. Overall, our results demonstrate that despite proximity to the Sahara and the Arabian Peninsula and the dominance of mineral dust in the aerosol sampled, existing mineral dust parameterizations alone would not skillfully represent the near-surface ns in the observed temperature regime (−6 to −25 ∘C). Future efforts to develop or improve representations of dust INPs at modest supercooling (≥-15 ∘C) would benefit from a characterization of the specific organic species associated with dust INPs. More generally, an improved understanding of the organic species associated with increased IN activity and their variability across dust source regions would directly inform efforts to determine whether ns-based parameterizations are appropriate for faithful representation of dust INPs in this sensitive temperature regime, whether region-specific parameterizations are required, or whether an alternative to the ns approach is necessary.

Pseudomonas syringae pv. syringae causes bacterial canker on Japanese quince (Chaenomeles japonica)

Japanese quince trees are grown as ornamental plants in Iran, in parks and in orchards close to stone fruit and pome fruit trees. Shoots of Japanese quince (Chaenomeles japonica) showing sunken brown canker symptoms were observed and collected near Sari, the center of Mazandaran province in the North of Iran, during the 2016 growing season. Gram negative bacteria isolated from symptomatic tissues were similar to Pseudomonas syringae pv. syringae (Pss) were pathogenic on Japanese quince and on quince (Cydonia oblonga) seedlings after artificial inoculation, and were re-isolated from diseased hosts. Phylogenetic tree construction using partial sequences of ITS and rpoD genes showed that the Japanese quince isolates were in the same clade as Pss strains. The isolates had ice nucleation activity, and the InaK gene was amplified successfully. According to the results of phenotypic and genotypic characteristics, genomic DNA fingerprinting using REP-PCR, BOX-PCR and IS50-PCR and isolation of total cell proteins, we conclude that Pss is the causal agent of canker of the Japanese quince trees. Therefore, Japanese quince is a new host for Pss causing bacterial canker on many different host plants.

Identification of Candidate Ice Nucleation Activity (INA) Genes in Fusarium avenaceum by Combining Phenotypic Characterization with Comparative Genomics and Transcriptomics.

Ice nucleation activity (INA) is the capacity of certain particles to catalyze ice formation at temperatures higher than the temperature at which pure water freezes. INA impacts the ratio of liquid to frozen cloud droplets and, therefore, the formation of precipitation and Earth’s radiative balance. Some Fusarium strains secrete ice-nucleating particles (INPs); they travel through the atmosphere and may thus contribute to these atmospheric processes. Fusarium INPs were previously found to consist of proteinaceous aggregates. Here, we determined that in F. avenaceum, the proteins forming these aggregates are smaller than 5 nm and INA is higher after growth at low temperatures and varies among strains. Leveraging these findings, we used comparative genomics and transcriptomics to identify candidate INA genes. Ten candidate INA genes that were predicted to encode secreted proteins were present only in the strains that produced the highest number of INPs. In total, 203 candidate INA genes coding for secreted proteins were induced at low temperatures. Among them, two genes predicted to encode hydrophobins stood out because hydrophobins are small, secreted proteins that form aggregates with amphipathic properties. We discuss the potential of the candidate genes to encode INA proteins and the next steps necessary to identify the molecular basis of INA in F. avenaceum.

Development and validation of a new cloud simulation experiment for lab-based aerosol-cloud studies.

The Aerosol Interaction and Dynamics in the Atmosphere (AIDA) cloud expansion chamber with a volume of 84m3 was extended for the small cloud expansion chamber AIDA mini (AIDAm) with a volume of 20 L. AIDAm is located in the cold room of AIDA and can perform automated ice-nucleation measurements over longer time periods of hours to days. AIDAm samples from the AIDA chamber, which acts as a reservoir of atmospheric aerosol types, which can slowly be modified by physical or chemical processes similar to those occurring in the atmosphere. AIDAm was validated for accurate ice-nucleation temperature control by measuring homogeneous freezing of pure water droplets at temperatures around -34 °C and for immersion freezing induced by dust aerosol particles in the temperature range between -20 and -30 °C. Further validation experiments at cirrus cloud temperatures of -45 °C revealed that AIDAm can distinguish between heterogeneous ice formation on mineral dust aerosols and homogeneous freezing of sulfuric acid solution particles. The contribution of homogeneous and heterogeneous ice formation processes to the ice-nucleation activity of coated dust particles was investigated in a 7 h long experiment, where solid dust particles were slowly coated with sulfuric acid. The continuous AIDAm measurements with a time resolution of 6minshowed a substantial suppression of the heterogeneous freezing phenomenon and an increasing role of homogeneous freezing while the coating amount was slowly increased. This experiment proved the capability of AIDAm to sensitively detect small changes in the ice-nucleation ability of aerosols, which undergo slow processing like chemical surface coating.

Silica as a Model Ice-Nucleating Particle to Study the Effects of Crystallinity, Porosity, and Low-Density Surface Functional Groups on Immersion Freezing

Aerosol particles can facilitate heterogeneous ice formation in the troposphere and stratosphere by acting as ice-nucleating particles, modulating cloud formation/dissipation, precipitation, and their microphysical properties. Heterogeneous ice nucleation is driven by ice embryo formation on the particle surface, which can be influenced by features of the surface such as crystallinity, surface structure, lattice structure, defects, and functional groups. To characterize the effect of crystallinity, pores, and surface functional groups toward ice nucleation, samples of comparable silica systems, specifically, quartz, ordered and nonordered porous amorphous silica samples with a range of pore sizes (2-11 nm), and nonporous functionalized silica spheres, were used as models for mineral dust aerosol particles. The ice nucleation activity of these samples was investigated by using an immersion freezing chamber. The results suggest that crystallinity has a larger effect than porosity on ice nucleation activity, as all of the porous silica samples investigated had lower onset freezing temperatures and lower ice nucleation activities than quartz. Our findings also suggest that pores alone are not sufficient to serve as effective active sites and need some additional chemical or physical property, like crystallinity, to nucleate ice in immersion mode freezing. The addition of a low density of organic functional groups to nonporous samples showed little enhancement compared to the inherent nucleation activity of silica with native surface hydroxyl groups. The density of functional groups investigated in this work suggests that a different arrangement of surface groups may be needed for enhanced immersion mode ice nucleation activity. In summary, crystallinity dictates the ice nucleation activity of silica samples rather than porosity or low-density surface functional groups. This work has broader implications regarding the climate impacts resulting from ice cloud formation.

Water-organizing motif continuity is critical for potent ice nucleation protein activity

Bacterial ice nucleation proteins (INPs) can cause frost damage to plants by nucleating ice formation at high sub-zero temperatures. Modeling of Pseudomonas borealis INP by AlphaFold suggests that the central domain of 65 tandem sixteen-residue repeats forms a beta-solenoid with arrays of outward-pointing threonines and tyrosines, which may organize water molecules into an ice-like pattern. Here we report that mutating some of these residues in a central segment of P. borealis INP, expressed in Escherichia coli, decreases ice nucleation activity more than the section’s deletion. Insertion of a bulky domain has the same effect, indicating that the continuity of the water-organizing repeats is critical for optimal activity. The ~10 C-terminal coils differ from the other 55 coils in being more basic and lacking water-organizing motifs; deletion of this region eliminates INP activity. We show through sequence modifications how arrays of conserved motifs form the large ice-nucleating surface required for potency.

Atomistic and coarse-grained simulations reveal increased ice nucleation activity on silver iodide surfaces in slit and wedge geometries

Abstract. Ice clouds can form at low and moderate supercooling through heterogeneous ice nucleation on atmospheric particles. Typically, the nucleation requires active sites with special chemical and physical properties, including surface topology and roughness. This paper investigates microscopic mechanisms of how combinations of confinement by the surface topology and lattice match induced by the surface properties can lead to enhanced ice nucleation. We perform molecular dynamics simulations using both atomistic and coarse-grained water models, at very low supercooling, to extensively study heterogeneous ice nucleation in slit-like and concave wedge structures of silver-terminated silver iodide (0001) surfaces. We find that ice nucleation is greatly enhanced by slit-like structures when the gap width is a near-integer multiple of the thickness of an ice bilayer. For wedge systems we also do not find a simple linear dependence between ice nucleation activity and the opening angle. Instead we observe strong enhancement in concave wedge systems with angles that match the orientations of ice lattice planes, highlighting the importance of structural matching for ice nucleation in confined geometries. While in the slit systems ice cannot grow out of the slit, some wedge systems show that ice readily grows out of the wedge. In addition, some wedge systems stabilize ice structures when heating the system above the thermodynamics melting point. In the context of atmospheric ice-nucleating particles, our results strongly support the experimental evidence for the importance of surface features such as cracks or pits functioning as active sites for ice nucleation at low supercooling.

Isolation and Identification of Ice Nucleation Active Fusarium Strains from Rapid Apple Declined Trees in Korea.

In biological particles such as Fusarium species, ice nucleation activity (INA) has been observed. Fusarium strains isolated from apple declined trees in Korea were identified with a multilocus sequence analysis using the tef1 and rpb1 genes. Droplet-freezing and tube-freezing assays were used to determine the INA of the strains, using Pseudomonas syringae pv. syringae KACC 21200 as a positive control and resulting in seven INA+ fungal strains that were identified as F. tricinctum (KNUF-21-F17, KNUF-21-F18, KNUF-21-F29, KNUF-21-F32, KNUF-21-F38, KNUF-21-F43, and KNUF-21-F44). The effect of Fusarium INA+ KNUF-21-F29 was compared to that of INA– strains on Chrysanthemum morifolium cv. Shinma explants. A higher callus formation and no-shoot formation were observed, suggesting that fungal INA could play a role in cold injuries and be a factor to consider in rapid apple decline. To the best of our knowledge, this is the first report of INA fungal strains isolated in Korea.

The ice-nucleating activity of African mineral dust in the Caribbean boundary layer

Abstract. African mineral dust is transported many thousands of kilometres from its source regions, and, because of its ability to nucleate ice, it plays a major role in cloud glaciation around the globe. The ice-nucleating activity of desert dust is influenced by its mineralogy, which varies substantially between source regions and across particle sizes. However, in models it is often assumed that the activity (expressed as active sites per unit surface area as a function of temperature) of atmospheric mineral dust is the same everywhere on the globe. Here, we find that the ice-nucleating activity of African desert dust sampled in the summertime marine boundary layer of Barbados (July and August 2017) is substantially lower than parameterizations based on soil from specific locations in the Sahara or dust sedimented from dust storms. We conclude that the activity of dust in Barbados' boundary layer is primarily defined by the low K-feldspar content of the dust, which is around 1 %. We propose that the dust we sampled in the Caribbean was from a region in western Africa (in and around the Sahel in Mauritania and Mali), which has a much lower feldspar content than other African sources across the Sahara and Sahel.

Ice nucleation active bacteria and its mitigation on tomato (Solanum lycopersicum)

Microbes play a vital role in ice nucleation, supporting bio-precipitation, and allowing plants to live in low-water environments. A field experiment was conducted during December 2018 with two phyllosphere microorganisms’ spraying viz., Pseudomonas aeruginosa and pink-pigmented facultative methylotrophs (PPFM) and under three moisture regimes (0.6, 0.8, 1.0 IW/CPE (irrigation water/ cumulative pan evaporation) ratio) on tomato (PKM 1) in Tamil Nadu Agricultural University, Coimbatore (Tamil Nadu). A laboratory experiment was conducted to confirm ice nucleation using two phyllosphere microorganisms’ P. aeruginosa and PPFM. The bioprecipitation impacts on tomatoes were assessed using a set of physiological parameters such as photosynthetic rate, average chlorophyll Index and the ice nucleation activity (INA) assessed using tube nucleation test and scanning electron microscope (SEM). The results showed that the mean photosynthetic rate of PPFM sprayed tomato (Solanum lycopersicum) (PKM 1) plants (40.7 μmol CO2/m2/s) at 10 DAS was significantly higher than P. aeruginosa sprayed plant (38.7 μmol CO2/m2/s) under different irrigation regimes. The average chlorophyll Index value of the P. aeruginosa sprayed tomato plants (58.1) was higher than PPFM sprayed plants (56.4). The tube nucleation tests were proved that ice crystallization induced by P. aeruginosa in super-cooled buffer at - 2 to -10°C while PPFM not catalyze the buffer even after 3hours. The scanning electron microscope (SEM) indicated the P. aeruginosa growth at the upper surface of the leaf and PPFM growth more at the lower surface of the leaf compared to without inoculation of microbes on leaves. Overall, the result revealed that P. aeruginosa may assist in ice nucleation activity that will help to make artificial rain in the near future.

Ice Nucleation Active bacteria in Mount Lawu forest, Indonesia: 2. Identification and characterization of ina gene bacteria isolated from lichens

Abstract. Arifin TN, Susilowati A, Sutarno. 2018. Ice Nucleation Active bacteria in Mount Lawu forest, Indonesia: 2. Identification and characterization of ina gene bacteria isolated from lichens. Asian J For 2: 39-46. Ice nucleation active (INA) bacteria can catalyze ice formation. Moreover, these bacteria cause frost injury in plants. This study aimed to determine the ina bacteria species of Mount Lawu forest, Java, Indonesia, based on the 16S rRNA gene and the characters of ice nucleation active gene coding of ina bacteria on lichens used ina gene primer. First, the isolates of ina bacteria were grown in NAG, and the DNA could be isolated. After that, genes coding for 16S rRNA and ina were amplified, and the amplification products were sequenced. Furthermore, the sequences were analyzed with BLAST program to know the similarity of bacteria species and ina gene characters. The result showed that the isolated ina bacteria had similarities with Pantoea, Pseudomonas, and Rahnella. The ina gene from N.2.1.B-13 isolate consists of 189 amino acids, dominated by a common amino acid found in other ina genes, like Alanine, Glycine, Tyrosine, Serine, Leucine, and Threonine. N terminal and C terminal are not found because ina gene from N.2.1.B-13 isolate is similar to the ice nucleation protein gene Pseudomonas borealis from bases 2947 bp to 3491 bp or 871 to 1060 amino acids.

Ice Nucleation Active bacteria in Mount Lawu forest, Indonesia: 1. Isolation and estimation of bacterial populations on lichens

Abstract. Fu’adah K, Sari SL, Susilowati A. 2017. Ice Nucleation Active bacteria in Mount Lawu forest, Indonesia: 1. Isolation and estimation of bacterial populations on lichens. Asian J For 1: 83-91. Ice Nucleation Active (INA) bacterias cause frost injury in plants. These bacteria are also predicted to play an important role in bioprecipitation, cloud formation, and rain. Most INA bacteria studies are conducted in subtropical areas. Therefore, studies on INA bacteria from tropical areas, especially Indonesia, need to be conducted. This study aimed to isolate and determine the number of INA bacteria in lichenes. samples of lichenes were taken from the hiking pathway of Cemoro Sewu, Mount Lawu forest, Java, Indonesia, at ±2.200, ±2.400, and ±2.600 m asl. INA bacteria were isolated by the spread plate method on the NA medium with 2.5 % glycerol and King’s B medium. A tube nucleation test determined ice nucleation activity. A multiple-tube nucleation test estimated the number of INA bacteria. The data were analyzed descriptively based on colony morphology, cell morphology, biochemical tests, and analysis of the numbers of INA bacteria. The result showed that 7 isolates from Parmelia sp. at an altitude of 2.532 m asl (station 2) have been known as INA bacterias. The number of INA bacteria in lichenes was 5 x 104/g, which was very low.

Structure and Protein-Protein Interactions of Ice Nucleation Proteins Drive Their Activity

Microbially-produced ice nucleating proteins (INpro) are unique molecular structures with the highest known catalytic efficiency for ice formation. Airborne microorganisms utilize these proteins to enhance their survival by reducing their atmospheric residence times. INpro also have critical environmental effects including impacts on the atmospheric water cycle, through their role in cloud and precipitation formation, as well as frost damage on crops. INpro are ubiquitously present in the atmosphere where they are emitted from diverse terrestrial and marine environments. Even though bacterial genes encoding INpro have been discovered and sequenced decades ago, the details of how the INpro molecular structure and oligomerization foster their unique ice-nucleation activity remain elusive. Using machine-learning based software AlphaFold 2 and trRosetta, we obtained and analysed the first ab initio structural models of full length and truncated versions of bacterial INpro. The modeling revealed a novel beta-helix structure of the INpro central repeat domain responsible for ice nucleation activity. This domain consists of repeated stacks of two beta strands connected by two sharp turns. One beta-strand is decorated with a TxT amino acid sequence motif and the other strand has an SxL[T/I] motif. The core formed between the stacked beta helix-pairs is unusually polar and very distinct from previous INpro models. Using synchrotron radiation circular dichroism, we validated the β-strand content of the central repeat domain in the model. Combining the structural model with functional studies of purified recombinant INpro, electron microscopy and modeling, we further demonstrate that the formation of dimers and higher-order oligomers is key to INpro activity. Using computational docking of the new INpro model based on rigid-body algorithms we could reproduce a previously proposed homodimer structure of the INpro CRD with an interface along a highly conserved tyrosine ladder and show that the dimer model agrees with our functional data. The parallel dimer structure creates a surface where the TxT motif of one monomer aligns with the SxL[T/I] motif of the other monomer widening the surface that interacts with water molecules and therefore enhancing the ice nucleation activity. This work presents a major advance in understanding the molecular foundation for bacterial ice-nucleation activity.

Brachypodium Antifreeze Protein Gene Products Inhibit Ice Recrystallisation, Attenuate Ice Nucleation, and Reduce Immune Response.

Antifreeze proteins (AFPs) from the model crop, Brachypodium distachyon, allow freeze survival and attenuate pathogen-mediated ice nucleation. Intriguingly, Brachypodium AFP genes encode two proteins, an autonomous AFP and a leucine-rich repeat (LRR). We present structural models which indicate that ice-binding motifs on the ~13 kDa AFPs can “spoil” nucleating arrays on the ~120 kDa bacterial ice nucleating proteins used to form ice at high sub-zero temperatures. These models are consistent with the experimentally demonstrated decreases in ice nucleating activity by lysates from wildtype compared to transgenic Brachypodium lines. Additionally, the expression of Brachypodium LRRs in transgenic Arabidopsis inhibited an immune response to pathogen flagella peptides (flg22). Structural models suggested that this was due to the affinity of the LRR domains to flg22. Overall, it is remarkable that the Brachypodium genes play multiple distinctive roles in connecting freeze survival and anti-pathogenic systems via their encoded proteins’ ability to adsorb to ice as well as to attenuate bacterial ice nucleation and the host immune response.

Preferential freezing avoidance localised in anthers and embryo sacs in wintering Daphne kamtschatica var. jezoensis flower buds visualised by magnetic resonance imaging.

To explore diversity in cold hardiness mechanisms, high resolution magnetic resonance imaging (MRI) was used to visualise freezing behaviours in wintering Daphne kamtschatica var. jezoensis flower buds, which have naked florets and no bud scales. MRI images showed that anthers remained stably supercooled to the range from -14 to -21°C or lower while most other tissues froze by -7°C. Freezing of some anthers detected in MRI images between -14 and -21°C corresponded with numerous low temperature exotherms and also with the 'all-or-nothing' type of anther injuries. In ovules/pistils, only embryo sacs remained supercooled at -7°C or lower, but slowly dehydrated during further cooling. Cryomicroscopic observation revealed ice formation in the cavities of calyx tubes and pistils but detected no ice in embryo sacs or in anthers. The distribution of ice nucleation activity in floral tissues corroborated the tissue freezing behaviours. Filaments likely work as the ice blocking barrier that prevents ice intrusion from extracellularly frozen calyx tubes to connecting unfrozen anthers. Unique freezing behaviours were demonstrated in Daphne flower buds: preferential freezing avoidance in male and female gametophytes and their surrounding tissues (by stable supercooling in anthers and by supercooling with slow dehydration in embryo sacs) while the remaining tissues tolerate extracellular freezing.

Effects of heterogeneous reaction with NO2 on ice nucleation activities of feldspar and Arizona Test Dust

Mineral dust is an important type of ice nucleating particles in the troposphere; however, the effects of heterogeneous reactions on ice nucleation (IN) activities of mineral dust remain to be elucidated. A droplet-freezing apparatus (Guangzhou Institute of Geochemistry Ice Nucleation Apparatus, GIGINA) was developed in this work to measure IN activities of atmospheric particles in the immersion freezing mode, and its performance was validated by a series of experimental characterizations. This apparatus was then employed to measure IN activities of feldspar and Arizona Test Dust (ATD) particles before and after heterogeneous reaction with NO2 (10±0.5 ppmv) at 40% relative humidity. The surface coverage of nitrate, θ(NO3−), increased to 3.1±0.2 for feldspar after reaction with NO2 for 6 hr, and meanwhile the active site density per unit surface area (ns) at -20°C was reduced from 92±5 to <1.0 cm−2 by about two orders of magnitude; however, no changes in nitrate content or IN activities were observed for further increase in reaction time (up to 24 hr). Both nitrate content and IN activities changed continuously with reaction time (up to 24 hr) for ATD particles; after reaction with NO2 for 24 hr, θ(NO3−) increased to 1.4±0.1 and ns at -20°C was reduced from 20±4 to 9.7±1.9 cm−2 by a factor of ∼2. Our work suggests that heterogeneous reaction with NO2, an abundant reactive nitrogen species in the troposphere, may significantly reduce IN activities of mineral dust in the immersion freezing mode.

Prevalence and characterization of Ice Nucleation Active (INA) bacteria from rainwater in Indonesia

BackgroundIce nucleation active (INA) bacteria are a group of microorganisms that can act as biological nucleator due to their ice nucleation protein property. Unfortunately, little is known about their prevalence and characteristics in tropical areas including Indonesia. Here, we monitor the presence of INA bacteria in rainwater and air samples collected from Jakarta, Tangerang and several areas in Western Java, Indonesia for one year. We further identify and characterize selected Class A of INA bacteria isolated from these areas.ResultsMost of the INA bacteria were isolated from rainwater samples collected during March–August 2010, particularly from Jakarta, Bandung, and Tangerang. A total of 1,902 bacterial isolates were recovered from these area. We found a limited number of bacterial isolates from air sampling. From ice nucleation activity assays, 101 INA isolates were found active as ice nucleator at a temperature above -10 °C. A large majority (73 isolates) of them are classified as Class C (active below -8 °C), followed by Class A (26 isolates; active at -2 to -5 °C) and Class B (two isolates; active at -5 to -8 °C). We sequenced the 16S rRNA gene of 18 Class A INA isolates and identified 15 isolates as Enterobacteriaceae, while the remaining three as Pseudomonadaceae. The vast majority of our Class A INA isolates were likely Pantoea spp. with several isolates were deduced as either Pseudomonas, Cronobacter, and Klebsiella. We found that these 18 Class A INA isolates had acquired resistance to antibiotics erythromycin and ampicillin, which are considered two critically important antibiotics.ConclusionsOur results showed that the prevalence of INA bacterial population varies across locations and seasons. Furthermore, our isolates were dominated by Class A and C INA bacteria. This study also cautions regarding the spread of antibiotic resistance among INA bacteria.

Laboratory studies of ice nucleation onto bare and internally mixed soot–sulfuric acid particles

Abstract. Soot particles are potential candidates for ice-nucleating particles in cirrus cloud formation, which is known to exert a net-warming effect on climate. Bare soot particles, generally hydrophobic and fractal ones, mainly exist near emission sources. Coated or internally mixed soot particles are more abundant in the atmosphere and have a higher probability of impacting cloud formation and climate. However, the ice nucleation ability of coated soot particles is not as well understood as that of freshly produced soot particles. In this laboratory study, two samples, a propane flame soot and a commercial carbon black, were used as atmospheric soot surrogates and coated with varying wt % of sulfuric acid (H2SO4). The ratio of coating material mass to the mass of bare soot particles was controlled and progressively increased from less than 5 wt % to over 100 wt %. Both bare and coated soot particle ice nucleation activities were investigated with a continuous-flow diffusion chamber operated at mixed-phase and cirrus cloud conditions. The mobility diameter and mass distribution of size-selected soot particles with/without H2SO4 coating were measured by a scanning mobility particle sizer and a centrifugal particle mass analyser running in parallel. The mixing state and morphology of soot particles were characterized by scanning electron microscopy and transmission electron microscopy. In addition, the evidence of the presence of H2SO4 on a coated soot particle surface is shown by energy-dispersive X-ray spectroscopy. Our study demonstrates that H2SO4 coatings suppress the ice nucleation activity of soot particles to varying degrees depending on the coating thickness, but in a non-linear fashion. Thin coatings causing pore filling in the soot aggregate inhibits pore condensation and freezing. Thick coatings promote particle ice activation via droplet homogeneous freezing. Overall, our findings reveal that H2SO4 coatings will suppress soot particle ice nucleation abilities in the cirrus cloud regime, having implications for the fate of soot particles with respect to cloud formation in the upper troposphere.

First Report of Bacterial Leaf Spot of Coriander Caused by Pseudomonas syringae pv. coriandricola in China.

Coriander (Coriandrum sativum L.) or Chinese parsley is a culinary herb with multiple medicinal effects that are widely used in cooking and traditional medicine. From September to November 2019, symptoms were observed in 2-month-old coriander plants from coriander fields in Lanzhou and Wenzhou, China. The disease developed rapidly under cold and wet climatic conditions, and the infection rate was almost 80% in open coriander fields. Typical symptoms on leaves included small, water-soaked blotches and irregular brown spots surrounding haloes; as the disease progressed, the spots coalesced into necrotic areas. Symptomatic leaf tissue was surface sterilized, macerated in sterile distilled water, and cultured on nutrient agar plates at 28 °C for 48 h (Koike and Bull, 2006). After incubation, six bacterial colonies, which were individually isolated from collected samples from two different areas, were selected for further study. Colonies on NA plate were small, round, raised, white to cream-colored, and had smooth margins. All bacterial isolates were gram-negative, rod-shaped and nonfluorescent on King's B medium. The bacteria were positive for levan production, Tween 80 hydrolysis, and tobacco hypersensitivity but negative for oxidase, potato slice rot test, arginine dihydrolase, ice nucleation activity, indole production and H2S production. The suspension of representative isolate for inoculating of plants was obtained from single colony on King's B medium for 2-3 days at 28 °C. DNA was extracted from bacterial suspensions of YS2003200102 cultured in 20 ml of King's B medium broth at 28 °C for 1 day. Extraction was performed with a TIANamp Bacterial DNA Kit (TIANGEN, China) according to the manufacturer's recommendations. The pathogen was confirmed by amplification and sequencing of the glyceraldehyde-3-phosphate dehydrogenase A (gapA) gene, the citrate synthase (gltA) gene, the DNA gyrase B (gyrB) gene and the RNA polymerase sigma factor 70 (rpoD) gene using gapA-For/gapA-Rev, gltA-For/gltA-Rev, gyrB-For/gryB-Rev, rpoD-For/rpoD-Rev primers, respectively (Popović et al., 2019). The sequences of the PCR products were deposited in GenBank with accession numbers MZ681931 (gapA), MZ681932 (gltA), MZ681933 (gyrB), and MZ681934 (rpoD). Phylogenetic analysis of multiple genes (Xu and Miller, 2013) was conducted with the maximum likelihood method using MEGA7. The sequences of our isolates and ten published sequences of P. syringae pv. coriandricola were clustered into one clade with a 100% confidence level. To confirm the pathogenicity of isolate YS2003200102, 2-month-old healthy coriander plants were inoculated by spraying the leaves with a bacterial suspension (108 CFU ml-1) at 28 °C incubation temperature and 70% relative humidity condition, and sterile distilled water was applied as a negative control treatment (Cazorla et al. 2005). Three replicates were conducted for every isolate, and each replicate included 6 coriander plants. After twelve days, only the inoculated leaves with bacterial suspension showed bacterial leaf spot resembling those observed on naturally infected coriander leaves. Cultures re-isolated from symptomatic leaves showed the same morphological characteristics and molecular traits as those initially isolated from infected leaves in the field. This bacterium was previously reported causing leaf spot of coriander in India and Spain (Gupta et al. 2013; Cazorla et al. 2005). To our knowledge, this is the first report of P. syringae pv. coriandricola causing leaf spot disease on coriander in China. Studies are needed on strategies to manage P. syringae pv. coriandricola in crops, because its prevalence may cause yield loss on coriander in China.