Effect of oxic and anoxic conditions on intracellular storage of polyhydroxyalkanoate and polyphosphate in Magnetospirillum magneticum strain AMB-1.

Magnetotactic bacteria (MTB) are microorganisms widely inhabiting the oxic-anoxic interface of aquatic environments. Beside biomineralizing magnetic nanocrystals, MTBs are able to sequester various chemical elements (e.g., carbon and phosphorus) for the biogenesis of intracellular granules, like polyhydroxyalkanoate (PHA) and polyphosphate (polyP), making them potentially important in biogeochemical cycling. Yet, the environmental controls of intracellular storage of carbon and phosphorus in MTB remain poorly understood. Here, we investigated the influence of oxic, anoxic and transient oxic-anoxic conditions on intracellular storage of PHA and polyP in Magnetospirillum magneticum strain AMB-1. In the incubations with oxygen, transmission electron microscopy revealed intercellular granules highly rich in carbon and phosphorus, which were further interpreted as PHA and polyP based on chemical and Energy-Dispersive X-ray spectroscopy analysis. Oxygen had a strong effect on PHA and polyP storage in AMB-1 cells, as PHA and polyP granules accounted for up to 47 ± 23% and 5.1 ± 1.7% of the cytoplasmic space, respectively, during continuous oxic conditions, while granules disappeared in anoxic incubations. Poly 3-hydroxybutyrate (PHB) and poly 3-hydroxyvalerate (PHV) accounted for 0.59 ± 0.66% and 0.0033 ± 0.0088% of dry cell weight, respectively, in anoxic incubations, while the values increased by a factor of 7 and 37 after oxygen was introduced. The results highlight a tight link between oxygen, carbon and phosphorus metabolisms in MTB, where favorable oxic growth conditions can lead to metabolic induction of polyP and PHA granule biogenesis.

Corrigendum: Effect of oxic and anoxic conditions on intracellular storage of polyhydroxyalkanoate and polyphosphate in Magnetospirillum magneticum strain AMB-1.

[This corrects the article DOI: 10.3389/fmicb.2023.1203805.].

Continuous Production of Biogenic Magnetite Nanoparticles by the Marine Bacterium Magnetovibrio blakemorei Strain MV-1T with a Nitrous Oxide Injection Strategy.

Magnetotactic bacteria (MTB) produce magnetosomes, which are membrane-embedded magnetic nanoparticles. Despite their technological applicability, the production of magnetite magnetosomes depends on the cultivation of MTB, which results in low yields. Thus, strategies for the large-scale cultivation of MTB need to be improved. Here, we describe a new approach for bioreactor cultivation of Magnetovibrio blakemorei strain MV-1T. Firstly, a fed-batch with a supplementation of iron source and N2O injection in 24-h pulses was established. After 120 h of cultivation, the production of magnetite reached 24.5 mg∙L-1. The maximum productivity (16.8 mg∙L-1∙day-1) was reached between 48 and 72 h. However, the productivity and mean number of magnetosomes per cell decreased after 72 h. Therefore, continuous culture in the chemostat was established. In the continuous process, magnetite production and productivity were 27.1 mg∙L-1 and 22.7 mg∙L-1∙day-1, respectively, at 120 h. This new approach prevented a decrease in magnetite production in comparison to the fed-batch strategy.

Biogeochemical fingerprinting of magnetotactic bacterial magnetite.

Biominerals are important archives of the presence of life and environmental processes in the geological record. However, ascribing a clear biogenic nature to minerals with nanometer-sized dimensions has proven challenging. Identifying hallmark features of biologically controlled mineralization is particularly important for the case of magnetite crystals, resembling those produced by magnetotactic bacteria (MTB), which have been used as evidence of early prokaryotic life on Earth and in meteorites. We show here that magnetite produced by MTB displays a clear coupled C-N signal that is absent in abiogenic and/or biomimetic (protein-mediated) nanometer-sized magnetite. We attribute the presence of this signal to intracrystalline organic components associated with proteins involved in magnetosome formation by MTB. These results demonstrate that we can assign a biogenic origin to nanometer-sized magnetite crystals, and potentially other biominerals of similar dimensions, using unique geochemical signatures directly measured at the nanoscale. This finding is significant for searching for the earliest presence of life in the Earth's geological record and prokaryotic life on other planets.

Complete Genome Sequence of Strain SS-5, a Magnetotactic Gammaproteobacterium Isolated from the Salton Sea, a Shallow, Saline, Endorheic Rift Lake Located on the San Andreas Fault in California.

We report the 3.7-Mb genome sequence of strain SS-5, a magnetotactic, sulfur-oxidizing rod and member of the family Chromatiaceae of the class Gammaproteobacteria, which biomineralizes membrane-bound, elongated, prismatic octahedral, magnetite nanocrystals. This genome sequence brings further diversity for understanding the origin and evolution of magnetotaxis and magnetosome biomineralization.

A rapid and simple preparation of amphotericin B-loaded bacterial magnetite nanoparticles.

Magnetotactic bacteria, which synthesize biological magnetite nanoparticles (BMs), are the main microbial source of magnetic nanomaterials. Although the use of BMs has been explored in vitro and in vivo for new anticancer formulations, targeted treatments of fungal and parasitic diseases would also benefit from biogenic magnetic nanoformulations. Due to the necessity of new formulations of amphotericin B, we developed a magnetic-nanoparticle based conjugate of this drug using bacterial magnetosomes. Different amphotericin B preparations were obtained using BMs extracted from Magnetovibrio blakemorei strain MV-1T as well as glutaraldehyde and poly-l-lysine as linking reagents. The highest capture efficiencies and drug loadings were achieved using 0.1‰ poly-l-lysine as the only linking agent (52.7 ± 2.1%, and 25.3 ± 1.9 µg per 100 µg, respectively) and 0.1‰ poly-l-lysine and glutaraldehyde 12.5% (45.0 ± 5.4%, and 21.6 ± 4.9 µg per 100 µg, respectively). Transmission electron microscopy and infrared spectroscopy analyses confirmed the association of amphotericin B to the BM surface. Moreover, controlled drug release from these nanoparticles was achieved by applying an alternating magnetic field. In this condition the release of amphotericin B in PBS increased approximately four-fold as compared to the release under standard conditions with no applied magnetic fields. Hence, the functionalization of BMs with amphotericin B produces stable nanoformulations with a controllable drug release profile, thus, enabling its potential in the treatment of fungal and parasitic diseases.

Expanding magnetic organelle biogenesis in the domain Bacteria.

BACKGROUND: The discovery of membrane-enclosed, metabolically functional organelles in Bacteria has transformed our understanding of the subcellular complexity of prokaryotic cells. Biomineralization of magnetic nanoparticles within magnetosomes by magnetotactic bacteria (MTB) is a fascinating example of prokaryotic organelles. Magnetosomes, as nano-sized magnetic sensors in MTB, facilitate cell navigation along the local geomagnetic field, a behaviour referred to as magnetotaxis or microbial magnetoreception. Recent discovery of novel MTB outside the traditionally recognized taxonomic lineages suggests that MTB diversity across the domain Bacteria are considerably underestimated, which limits understanding of the taxonomic distribution and evolutionary origin of magnetosome organelle biogenesis. RESULTS: Here, we perform the most comprehensive metagenomic analysis available of MTB communities and reconstruct metagenome-assembled MTB genomes from diverse ecosystems. Discovery of MTB in acidic peatland soils suggests widespread MTB occurrence in waterlogged soils in addition to subaqueous sediments and water bodies. A total of 168 MTB draft genomes have been reconstructed, which represent nearly a 3-fold increase over the number currently available and more than double the known MTB species at the genome level. Phylogenomic analysis reveals that these genomes belong to 13 Bacterial phyla, six of which were previously not known to include MTB. These findings indicate a much wider taxonomic distribution of magnetosome organelle biogenesis across the domain Bacteria than previously thought. Comparative genome analysis reveals a vast diversity of magnetosome gene clusters involved in magnetosomal biogenesis in terms of gene content and synteny residing in distinct taxonomic lineages. Phylogenetic analyses of core magnetosome proteins in this largest available and taxonomically diverse dataset support an unexpectedly early evolutionary origin of magnetosome biomineralization, likely ancestral to the origin of the domain Bacteria. CONCLUSIONS: These findings expand the taxonomic and phylogenetic diversity of MTB across the domain Bacteria and shed new light on the origin and evolution of microbial magnetoreception. Potential biogenesis of the magnetosome organelle in the close descendants of the last bacterial common ancestor has important implications for our understanding of the evolutionary history of bacterial cellular complexity and emphasizes the biological significance of the magnetosome organelle. Video Abstract.

In Vivo Biosynthesis of Inorganic Nanomaterials Using Eukaryotes-A Review.

Bionanotechnology, the use of biological resources to produce novel, valuable nanomaterials, has witnessed tremendous developments over the past two decades. This eco-friendly and sustainable approach enables the synthesis of numerous, diverse types of useful nanomaterials for many medical, commercial, and scientific applications. Countless reviews describing the biosynthesis of nanomaterials have been published. However, to the best of our knowledge, no review has been exclusively focused on the in vivo biosynthesis of inorganic nanomaterials. Therefore, the present review is dedicated to filling this gap by describing the many different facets of the in vivo biosynthesis of nanoparticles (NPs) using living eukaryotic cells and organisms-more specifically, live plants and living biomass of several species of microalgae, yeast, fungus, mammalian cells, and animals. It also highlights the strengths and weaknesses of the synthesis methodologies and the NP characteristics, bio-applications, and proposed synthesis mechanisms. This comprehensive review also brings attention to enabling a better understanding between the living organisms themselves and the synthesis conditions that allow their exploitation as nanobiotechnological production platforms as these might serve as a robust resource to boost and expand the bio-production and use of desirable, functional inorganic nanomaterials.

Insight on thermal stability of magnetite magnetosomes: implications for the fossil record and biotechnology.

Magnetosomes are intracellular magnetic nanocrystals composed of magnetite (Fe3O4) or greigite (Fe3S4), enveloped by a lipid bilayer membrane, produced by magnetotactic bacteria. Because of the stability of these structures in certain environments after cell death and lysis, magnetosome magnetite crystals contribute to the magnetization of sediments as well as providing a fossil record of ancient microbial ecosystems. The persistence or changes of the chemical and magnetic features of magnetosomes under certain conditions in different environments are important factors in biotechnology and paleomagnetism. Here we evaluated the thermal stability of magnetosomes in a temperature range between 150 and 500 °C subjected to oxidizing conditions by using in situ scanning transmission electron microscopy. Results showed that magnetosomes are stable and structurally and chemically unaffected at temperatures up to 300 °C. Interestingly, the membrane of magnetosomes was still observable after heating the samples to 300 °C. When heated between 300 °C and 500 °C cavity formation in the crystals was observed most probably associated to the partial transformation of magnetite into maghemite due to the Kirkendall effect at the nanoscale. This study provides some insight into the stability of magnetosomes in specific environments over geological periods and offers novel tools to investigate biogenic nanomaterials.

Complete Genome Sequence of Strain BW-2, a Magnetotactic Gammaproteobacterium in the Family Ectothiorhodospiraceae, Isolated from a Brackish Spring in Death Valley, California.

We report the complete 4.1-Mb genome sequence of strain BW-2, a magnetotactic, sulfur-oxidizing rod, belonging to the family Ectothiorhodospiraceae of the class Gammaproteobacteria, that biomineralizes membrane-bounded magnetite nanocrystals in its magnetosomes. This genome sequence, in comparison with those of other magnetotactic bacteria, is essential for understanding the origin and evolution of magnetotaxis and magnetosome biomineralization.

On the origin of microbial magnetoreception.

A broad range of organisms, from prokaryotes to higher animals, have the ability to sense and utilize Earth's geomagnetic field-a behavior known as magnetoreception. Although our knowledge of the physiological mechanisms of magnetoreception has increased substantially over recent decades, the origin of this behavior remains a fundamental question in evolutionary biology. Despite this, there is growing evidence that magnetic iron mineral biosynthesis by prokaryotes may represent the earliest form of biogenic magnetic sensors on Earth. Here, we integrate new data from microbiology, geology and nanotechnology, and propose that initial biomineralization of intracellular iron nanoparticles in early life evolved as a mechanism for mitigating the toxicity of reactive oxygen species (ROS), as ultraviolet radiation and free-iron-generated ROS would have been a major environmental challenge for life on early Earth. This iron-based system could have later been co-opted as a magnetic sensor for magnetoreception in microorganisms, suggesting an origin of microbial magnetoreception as the result of the evolutionary process of exaptation.

Magnetosome magnetite biomineralization in a flagellated protist: evidence for an early evolutionary origin for magnetoreception in eukaryotes.

The most well-recognized magnetoreception behaviour is that of the magnetotactic bacteria (MTB), which synthesize membrane-bounded magnetic nanocrystals called magnetosomes via a biologically controlled process. The magnetic minerals identified in prokaryotic magnetosomes are magnetite (Fe3 O4 ) and greigite (Fe3 S4 ). Magnetosome crystals, regardless of composition, have consistent, species-specific morphologies and single-domain size range. Because of these features, magnetosome magnetite crystals possess specific properties in comparison to abiotic, chemically synthesized magnetite. Despite numerous discoveries regarding MTB phylogeny over the last decades, this diversity is still considered underestimated. Characterization of magnetotactic microorganisms is important as it might provide insights into the origin and establishment of magnetoreception in general, including eukaryotes. Here, we describe the magnetotactic behaviour and characterize the magnetosomes from a flagellated protist using culture-independent methods. Results strongly suggest that, unlike previously described magnetotactic protists, this flagellate is capable of biomineralizing its own anisotropic magnetite magnetosomes, which are aligned in complex aggregations of multiple chains within the cell. This organism has a similar response to magnetic field inversions as MTB. Therefore, this eukaryotic species might represent an early origin of magnetoreception based on magnetite biomineralization. It should add to the definition of parameters and criteria to classify biogenic magnetite in the fossil record.

Magnetosome Gene Duplication as an Important Driver in the Evolution of Magnetotaxis in the Alphaproteobacteria.

The evolution of microbial magnetoreception (or magnetotaxis) is of great interest in the fields of microbiology, evolutionary biology, biophysics, geomicrobiology, and geochemistry. Current genomic data from magnetotactic bacteria (MTB), the only prokaryotes known to be capable of sensing the Earth's geomagnetic field, suggests an ancient origin of magnetotaxis in the domain Bacteria Vertical inheritance, followed by multiple independent magnetosome gene cluster loss, is considered to be one of the major forces that drove the evolution of magnetotaxis at or above the class or phylum level, although the evolutionary trajectories at lower taxonomic ranks (e.g., within the class level) remain largely unstudied. Here we report the isolation, cultivation, and sequencing of a novel magnetotactic spirillum belonging to the genus Terasakiella (Terasakiella sp. strain SH-1) within the class Alphaproteobacteria The complete genome sequence of Terasakiella sp. strain SH-1 revealed an unexpected duplication event of magnetosome genes within the mamAB operon, a group of genes essential for magnetosome biomineralization and magnetotaxis. Intriguingly, further comparative genomic analysis suggests that the duplication of mamAB genes is a common feature in the genomes of alphaproteobacterial MTB. Taken together, with the additional finding that gene duplication appears to have also occurred in some magnetotactic members of the Deltaproteobacteria, our results indicate that gene duplication plays an important role in the evolution of magnetotaxis in the Alphaproteobacteria and perhaps the domain Bacteria IMPORTANCE A diversity of organisms can sense the geomagnetic field for the purpose of navigation. Magnetotactic bacteria are the most primitive magnetism-sensing organisms known thus far and represent an excellent model system for the study of the origin, evolution, and mechanism of microbial magnetoreception (or magnetotaxis). The present study is the first report focused on magnetosome gene cluster duplication in the Alphaproteobacteria, which suggests the important role of gene duplication in the evolution of magnetotaxis in the Alphaproteobacteria and perhaps the domain Bacteria A novel scenario for the evolution of magnetotaxis in the Alphaproteobacteria is proposed and may provide new insights into evolution of magnetoreception of higher species.

Misalignment between the magnetic dipole moment and the cell axis in the magnetotactic bacterium Magnetospirillum magneticum AMB-1.

While most quantitative studies of the motion of magnetotactic bacteria rely on the premise that the cells' magnetic dipole moment is aligned with their direction of motility, this assumption has so far rarely been challenged. Here we use phase contrast microscopy to detect the rotational diffusion of non-motile cells of Magnetospirillum magneticum AMB-1 around their magnetic moment, showing that in this species the magnetic dipole moment is, in fact, not exactly aligned with the cell body axis. From the cell rotational trajectories, we are able to infer the misalignment between cell magnetic moment and body axis with a precision of better than 1°, showing that it is, on average, 6°, and can be as high as 20°. We propose a method to correct for this misalignment, and perform a non-biased measurement of the magnetic moment of single cells based on the analysis of their orientation distribution. Using this correction, we show that magnetic moment strongly correlates with cell length. The existence of a range of misalignments between magnetic moment and cell axis in a population implies that the orientation and trajectories of magnetotactic bacteria placed in external magnetic fields is more complex than generally assumed, and might show some important cell-to-cell differences.

Uptake and persistence of bacterial magnetite magnetosomes in a mammalian cell line: Implications for medical and biotechnological applications.

Magnetotactic bacteria biomineralize intracellular magnetic nanocrystals surrounded by a lipid bilayer called magnetosomes. Due to their unique characteristics, magnetite magnetosomes are promising tools in Biomedicine. However, the uptake, persistence, and accumulation of magnetosomes within mammalian cells have not been well studied. Here, the endocytic pathway of magnetite magnetosomes and their effects on human cervix epithelial (HeLa) cells were studied by electron microscopy and high spatial resolution nano-analysis techniques. Transmission electron microscopy of HeLa cells after incubation with purified magnetosomes showed the presence of magnetic nanoparticles inside or outside endosomes within the cell, which suggests different modes of internalization, and that these structures persisted beyond 120 h after internalization. High-resolution transmission electron microscopy and electron energy loss spectra of internalized magnetosome crystals showed no structural or chemical changes in these structures. Although crystal morphology was preserved, iron oxide crystalline particles of approximately 5 nm near internalized magnetosomes suggests that minor degradation of the original mineral structures might occur. Cytotoxicity and microscopy analysis showed that magnetosomes did not result in any apparent effect on HeLa cells viability or morphology. Based on our results, magnetosomes have significant biocompatibility with mammalian cells and thus have great potential in medical, biotechnological applications.

Growing Magnetotactic Bacteria of the Genus Magnetospirillum: Strains MSR-1, AMB-1 and MS-1.

Magnetotactic bacteria are Gram-negative, motile, mainly aquatic prokaryotes ubiquitous in freshwater and marine habitats. They are characterized by their ability to biomineralize magnetosomes, which are magnetic nanometer-sized crystals of magnetite (Fe3O4) or greigite (Fe3S4) surrounded by a lipid bilayer membrane, within their cytoplasm. For most known magnetotactic bacteria, magnetosomes are assembled in chains inside the cytoplasm, thereby conferring a permanent magnetic dipole moment to the cells and causing them to align passively with external magnetic fields. Because of these specific features, magnetotactic bacteria have a great potential for commercial and medical applications. However, most species are microaerophilic and have specific O2 concentration requirements, making them more difficult to grow routinely than many other bacteria such as Escherichia coli. Here we present detailed protocols for growing three of the most widely studied strains of magnetotactic bacteria, all belonging to the genus Magnetospirillum. These methods allow for precise control of the O2 concentration made available to the bacteria, in order to ensure that they grow normally and synthesize magnetosomes. Growing magnetotactic bacteria for further studies using these procedures does not require the experimentalist to be an expert in microbiology. The general methods presented in this article may also be used to isolate and culture other magnetotactic bacteria, although it is likely that growth media chemical composition will need to be modified.

Variation in airborne fungal spore concentrations among five monitoring locations in a desert urban environment.

Fungal spores are biological particles that are ubiquitous in the outdoor air. Spores of several very common fungal species are known allergens, with the potential to cause respiratory illnesses by exacerbating asthma and allergic rhinitis. The National Allergy Bureau typically has one monitoring station established per city to determine fungal spore counts for an entire metropolitan area. However, variations in fungal spore concentrations could occur among different locations. The objective of this study was to measure and compare airborne fungal spore concentrations in five locations in Las Vegas for the year 2015 to determine if there are differences among microenvironments in the city. Twenty-four-hour or 7-day air samples were collected from five sites across the Las Vegas Valley. Samples were analyzed with a light microscope for fungal spores and counts were converted to concentrations of spores per volume of air. Mixed-model methods were used to evaluate mean differences. Results showed that smuts (basidiomycetes) were the dominant spore type for all five sites during the spring season. Cladosporium species were responsible for the second most dominant spore type with the highest concentrations occurring during the summer and fall months. Results obtained from the five stations established in Las Vegas show that there are important variations among the sites regarding fungal spore concentrations. The data suggest that more sites and additional monitoring of outdoor allergens are needed to provide information necessary to inform the community of outdoor air quality conditions and their potential effects on public health. This study presents new outdoor fungal spore data for the southwest region of the USA, focused in the Las Vegas Valley.

Applications of Magnetotactic Bacteria, Magnetosomes and Magnetosome Crystals in Biotechnology and Nanotechnology: Mini-Review.

Magnetotactic bacteria (MTB) biomineralize magnetosomes, which are defined as intracellular nanocrystals of the magnetic minerals magnetite (Fe3O4) or greigite (Fe3S4) enveloped by a phospholipid bilayer membrane. The synthesis of magnetosomes is controlled by a specific set of genes that encode proteins, some of which are exclusively found in the magnetosome membrane in the cell. Over the past several decades, interest in nanoscale technology (nanotechnology) and biotechnology has increased significantly due to the development and establishment of new commercial, medical and scientific processes and applications that utilize nanomaterials, some of which are biologically derived. One excellent example of a biological nanomaterial that is showing great promise for use in a large number of commercial and medical applications are bacterial magnetite magnetosomes. Unlike chemically-synthesized magnetite nanoparticles, magnetosome magnetite crystals are stable single-magnetic domains and are thus permanently magnetic at ambient temperature, are of high chemical purity, and display a narrow size range and consistent crystal morphology. These physical/chemical features are important in their use in biotechnological and other applications. Applications utilizing magnetite-producing MTB, magnetite magnetosomes and/or magnetosome magnetite crystals include and/or involve bioremediation, cell separation, DNA/antigen recovery or detection, drug delivery, enzyme immobilization, magnetic hyperthermia and contrast enhancement of magnetic resonance imaging. Metric analysis using Scopus and Web of Science databases from 2003 to 2018 showed that applied research involving magnetite from MTB in some form has been focused mainly in biomedical applications, particularly in magnetic hyperthermia and drug delivery.

Association of magnetotactic multicellular prokaryotes with Pseudoalteromonas species in a natural lagoon environment.

Magnetotactic bacteria, for the most part, are free-living, motile, unicellular prokaryotes that inhabit almost all marine and freshwater environments. One notable exception to the unicellular mode, however, are the magnetotactic multicellular prokaryotes. These morphologically unique prokaryotes (e.g., Candidatus Magnetoglobus multicellularis) are motile aggregates of 20-40 genetically identical, Gram-negative cells organised as a sphere (or ovoid in shape) and only motile as a unit. No specific close physical association between magnetotactic bacteria and non-magnetotactic microorganisms has ever been reported. Here, using culture-independent approaches, we show an unusual association between the spherical magnetotactic multicellular prokaryote Ca. Magnetoglobus multicellularis and Pseudoalteromonas species in environmental sediment and water samples collected from the Araruama Lagoon in Brazil. Cells of Pseudoalteromonas species were observed to be physically attached to the surface and, notably, even in the intercellular space of these spherical magnetotactic multicellular prokaryotes. An attempt to correlate the frequency of association between Pseudoalteromonas and magnetotactic multicellular prokaryotes with sediment depth was made but only a slight decrease in the number of Pseudoalteromonas cells per magnetotactic multicellular prokaryote was observed with increasing depth. Similar observations were made with magnetotactic multicellular prokaryotes from another Brazilian Lagoon (Rodrigo de Freitas) and the putative symbiont/parasite was detected. Although our results suggest some sort of specificity in the relationship between these prokaryotes, the precise nature of this association remains unclear.

Variation in airborne pollen concentrations among five monitoring locations in a desert urban environment.

The urbanization of the Las Vegas Valley has transformed this part of the Mohave Desert into a green oasis by introducing many non-native plant species, some of which are allergenic. Typically, one monitoring station is established per city to obtain pollen counts for an entire metropolitan area. However, variations in pollen concentrations could occur among different microenvironments. The objective of this study is to measure and compare pollen concentrations in five locations in Las Vegas to determine if there are significant differences between microenvironments within the city. Air samples were collected from five sites across the Las Vegas Valley over a 1-year period. Prepared slides were analyzed with a light microscope for pollen grains and converted into airborne pollen concentrations. Mixed model methods were used to determine mean differences. Tree pollen was the greatest contributor to the annual average airborne pollen concentrations (130 grains/m3) compared to weeds (6 grains/m3) and grass (3 grains/m3). The highest peak occurred in March 2016 (9589 total grains/m3). There were several differences among sites with respect to concentrations of individual tree species and for total weed and grass concentrations. We observed significant variations in concentration and composition among the five pollen collection stations that were established across the Las Vegas Valley. This study presented new outdoor pollen data for the southwest region of the USA, focused in Las Vegas. The results indicate that more sites and comprehensive monitoring of outdoor allergens are needed to provide accurate information to the community about outdoor air quality conditions.