Culture-Independent Characterization of the Microbiota of the Ant Lion Myrmeleon mobilis (Neuroptera: Myrmeleontidae)

Ant lions are insect larvae that feed on the liquefied internal components of insect prey. Prey capture is assisted by the injection of toxins that are reportedly derived from both the insect and bacterial symbionts. These larvae display interesting gut physiology where the midgut is not connected to the hindgut, preventing elimination of solid waste until adulthood. The presence of a discontinuous gut and the potential involvement of bacteria in prey paralyzation suggest an interesting microbial role in ant lion biology; however, the ant lion microbiota has not been described in detail. We therefore performed culture-independent 16S rRNA gene sequence analysis of the bacteria associated with tissues of an ant lion, Myrmeleon mobilis. All 222 sequences were identified as Proteobacteria and could be subdivided into two main groups, the α-Proteobacteria with similarity to Wolbachia spp. (75 clones) and the γ-Proteobacteria with similarity to the family Enterobacteriaceae (144 clones). The Enterobacteriaceae-like 16S rRNA gene sequences were most commonly isolated from gut tissue, and Wolbachia-like sequences were predominant in the head and body tissue. Fluorescence in situ hybridization analyses supported the localization of enterics to gut tissue and Wolbachia to nongut tissue. The diversity of sequences isolated from freshly caught, laboratory-fed, and laboratory-starved ant lions were qualitatively similar, although the libraries from each treatment were significantly different (P = 0.05). These results represent the first culture-independent analysis of the microbiota associated with a discontinuous insect gut and suggest that the ant lion microbial community is relatively simple, which may be a reflection of the diet and gut physiology of these insects.     

Characterization of the Fecal Microbiota Using High-Throughput Sequencing Reveals a Stable Microbial Community during Storage

The handling and treatment of biological samples is critical when characterizing the composition of the intestinal microbiota between different ecological niches or diseases. Specifically, exposure of fecal samples to room temperature or long term storage in deep freezing conditions may alter the composition of the microbiota. Thus, we stored fecal samples at room temperature and monitored the stability of the microbiota over twenty four hours. We also investigated the stability of the microbiota in fecal samples during a six month storage period at −80°C. As the stability of the fecal microbiota may be affected by intestinal disease, we analyzed two healthy controls and two patients with irritable bowel syndrome (IBS). We used high-throughput pyrosequencing of the 16S rRNA gene to characterize the microbiota in fecal samples stored at room temperature or −80°C at six and seven time points, respectively. The composition of microbial communities in IBS patients and healthy controls were determined and compared using the Quantitative Insights Into Microbial Ecology (QIIME) pipeline. The composition of the microbiota in fecal samples stored for different lengths of time at room temperature or −80°C clustered strongly based on the host each sample originated from. Our data demonstrates that fecal samples exposed to room or deep freezing temperatures for up to twenty four hours and six months, respectively, exhibit a microbial composition and diversity that shares more identity with its host of origin than any other sample.     

Ingested Blood Contributes to the Specificity of the Symbiosis of Aeromonas veronii Biovar Sobria and Hirudo medicinalis, the Medicinal Leech

Hirudo medicinalis, the medicinal leech, usually carries in its digestive tract a pure culture of Aeromonas veronii bv. sobria. Such specificity is unusual for digestive tracts that are normally colonized by a complex microbial consortium. Important questions for the symbiotic interaction and for the medical application after microvascular surgery are whether other bacteria can proliferate or at least persist in the digestive tract of H. medicinalis and what factors contribute to the reported specificity. Using a colonization assay, we were able to compare experimentally the ability of clinical isolates and of a symbiotic strain to colonize H. medicinalis. The symbiotic A. veronii bv. sobria strain proliferated well and persisted for at least 7 days inside the digestive tract. In contrast, the proliferation of Pseudomonas aeruginosa and Staphylococcus aureus was inhibited inside the animal compared to growth in the in vitro control, indicating that the ingested blood was modified within the digestive tract. However, both strains were able to persist in the digestive tract for at least 7 days. For an Escherichia coli strain, the viable counts decreased approximately 1,000-fold within 42 h. The decrease of viable E. coli could be prevented by interfering with the activation of the membrane-attack complex of the complement system that is present in blood. This suggests that the membrane-attack complex remained active inside H. medicinalis and prevented the proliferation of sensitive bacteria. Thus, antimicrobial properties of the ingested vertebrate blood contribute to the specificity of the A. veronii-H. medicinalis symbiosis, in addition to modifications of the blood inside the digestive tract of H. medicinalis.     

Culture-Independent Analysis of Gut Bacteria: the Pig Gastrointestinal Tract Microbiota Revisited

The phylogenetic diversity of the intestinal bacterial community in pigs was studied by comparative 16S ribosomal DNA (rDNA) sequence analysis. Samples were collected from a total of 24 pigs representing a variety of diets, ages, and herd health status. A library comprising 4,270 cloned 16S rDNA sequences obtained directly by PCR from 52 samples of either the ileum, the cecum, or the colon was constructed. In total, 375 phylotypes were identified using a 97% similarity criterion. Three hundred nine of the phylotypes (83%) had a <97% sequence similarity to any sequences in the database and may represent yet-uncharacterized bacterial genera or species. The phylotypes were affiliated with 13 major phylogenetic lineages. Three hundred four phylotypes (81%) belonged to the low-G+C gram-positive division, and 42 phylotypes (11.2%) were affiliated with the Bacteroides and Prevotella group. Four clusters of phylotypes branching off deeply within the low-G+C gram-positive bacteria and one in the Mycoplasma without any cultured representatives were found. The coverage of all the samples was 97.2%. The relative abundance of the clones approximated a lognormal distribution; however, the phylotypes detected and their abundance varied between two libraries from the same sample. The results document that the intestinal microbial community is very complex and that the majority of the bacterial species colonizing the gastrointestinal tract in pigs have not been characterized.     

Modeling a Neural Oscillator that Paces Heartbeat in the Medicinal Leech

SYNOPSIS. Heartbeat in the medicinal leech is paced by a neuraloscillator comprising two elemental oscillators whose activityis coordinated intersegmental coordinating fibers. The elementaloscillators each consist of a bilateral pair of heart interneuronslinked by reciprocal inhibitory synapses. The activity cycleof each elemental oscillator consists of alternating burstsof action potentials (plateau/burst phase) and periods inhibition(inactive phase). Oscillation ensues in the reciprocally inhibitorypairs because each neuron is able to escape from the inhibitionits contralateral partner and thus move on to the plateau/burstphase. We have identified and described membrane currents thatcontribute to oscillation and studied graded synaptic transmissionbetween the neurons, using discontinuous current clamp and switchingsingle electrode voltage clamp techniques. A hyperpolarization-activatedinward current, Ih, plays a major role in escape from inhibition,and Ca²⁺ currents produce plateau potentials that support burstformation and mediate graded synaptic transmission. To consolidate our knowledge and guide future research, we haveconstructed a first generation computer model of a neural oscillatorbased on reciprocal inhibition, using Hodgkin-Huxley equationsand a synaptic transfer model, derived from our biophysicalstudies, with Nodus software (De Schutter, 1989). This modelhas confirmed an important role for I_h in sustaining oscillationand has implicated a similarly important role for outward currents(particularly I_A), which remain to be studied. Neural oscillatorsbased on reciprocal inhibition appear to be ubiquitous, andour studies, biophysical and computational, provide insightsinto how they may operate.     

Localized RNAi and Ectopic Gene Expression in the Medicinal Leech

In this video, we show the use of a pneumatic capillary gun for the accurate biolistic delivery of reagents into live tissue. We use the procedure to perturb gene expression patterns in selected segments of leech embryos, leaving the untreated segments as internal controls.The pneumatic capillary gun can be used to reach internal layers of cells at early stages of development without opening the specimen. As a method for localized introduction of substances into living tissues, the biolistic delivery with the gun has several advantages: it is fast, contact-free and non-destructive. In addition, a single capillary gun can be used for independent delivery of different substances. The delivery region can have lateral dimensions of ~50-150 µm and extends over ~15 µm around the mean penetration depth, which is adjustable between 0 and 50 µm. This delivery has the advantage of being able to target a limited number of cells in a selected location intermediate between single cell knock down by microinjection and systemic knockdown through extracellular injections or by means of genetic approaches.For knocking down or knocking in the expression of the axon guidance molecule Netrin, which is naturally expressed by some central neurons and in the ventral body wall, but not the dorsal domain, we deliver molecules of dsRNA or plasmid-DNA into the body wall and central ganglia. This procedure includes the following steps: (i) preparation of the experimental setup for a specific assay (adjusting the accelerating pressure), (ii) coating the particles with molecules of dsRNA or DNA, (iii) loading the coated particles into the gun, up to two reagents in one assay, (iv) preparing the animals for the particle delivery, (v) delivery of coated particles into the target tissue (body wall or ganglia), and (vi) processing the embryos (immunostaining, immunohistochemistry and neuronal labeling) to visualize the results, usually 2 to 3 days after the delivery.When the particles were coated with netrin dsRNA, they caused clearly visible knock-down of netrin expression that only occurred in cells containing particles (usually, 1-2 particles per cell). Particles coated with a plasmid encoding EGFP induced fluorescence in neuronal cells when they stopped in their nuclei.Download video file.^{(116M, mov)}     

The External Structure and Distribution of Sensilla in the Medicinal Leech

Abstract The body of Hirudo medicinalis consists of 32 segments. The quinquannulate midbody segments 3–18 bear 14 sensilla on the central (neural) annulus. Elsewhere segments are represented by fewer than five annuli but sensilla are retained on the neural annulus. From neural sensilla protrude S hairs (cilia from uniciliate cells) which are thought to detect water currents. Two categories of multiciliate cell, of unknown function, are also present within neural sensilla; (i) grouped cilia extending beyond the cuticle (G hairs), and (ii) cilia which radiate out beneath the cuticle. Studies of the entire external surface of leech body wall with a scanning electron microscope revealed the presence of large numbers of sensilla on every annulus including the neural annuli. These sensilla lack S hairs. Our results show: (i) annular sensilla have a significantly smaller surface area than neural sensilla (p < 0.001 Mann-Whitney test, 2 tailed), (ii) The position and number of the small sensilla varies from annulus to annulus, segment to segment and individual to individual, (iii) Significantly higher numbers of small sensilla were found where non-neural annuli remain single and have not undergone a further division during development as happens in the quinquannulate segments. The data suggest that small sensilla continue to be added during the adult life of the leech.     

Heritability of Reproductive Traits in the Medicinal Leech Hirudo medicinalis L.

The phenotype variability and inheritance of reproductive traits were investigated in the medicinal leech. Distribution parameters were determined for the following traits: batch size (X¯ = 4.3 ± 0.2, = 1.7, CV = 40%, As = 0.23 ± 0.25, Ex = 0.19 ± 0.51), number of juveniles in a cocoon ( X¯= 10.9 ± 0.3, = 4.6, CV = 42%, As = 0.31 ± 0.15, Ex = 0.23 ± 0.30), and juvenile weight ( X¯= 32.0 ± 0.3, = 14.9, CV = 47%, As = 1.38 ± 0.05, Ex = 3.32 ± 0.11). A nonlinear negative correlation between the number of juveniles in a cocoon and their weight was found (correlation ratio R= 0.86). It was shown that the environmental variance dominated over the genotypic one in the structure of phenotypic variance of the traits studied. The genetic variability is determined mainly by additive gene interactions and, to a small extent, intralocus dominance. The narrow-sense heritability, h ², for batch size was 0.35–0.40; for the number of juveniles in a cocoon, 0.35; for juvenile weight, 0.42.     

Novel Role for Aeromonas jandaei as a Digestive Tract Symbiont of the North American Medicinal Leech

The gut bacteria of the North American medicinal leech, Macrobdella decora, were characterized. Biochemical tests and DNA sequences indicated that Aeromonas jandaei is the dominant culturable symbiont in leeches from a broad geographic area. In this work we identified a new habitat for A. jandaei, and here we suggest that there is unexpected specificity between leeches and Aeromonas species.     

Culture-Independent Characterization of Archaeal Biodiversity in Swine Confinement Building Bioaerosols

It was previously demonstrated that microbial communities of pig manure were composed of both bacteria and archaea. Recent studies have shown that bacteria are aerosolized from pig manure, but none have ever focused on the airborne archaeal burden. We sought here to develop and apply molecular ecology approaches to thoroughly characterize airborne archaea from swine confinement buildings (SCBs). Eight swine operations were visited, twice in winter and once during summer. Institute of Occupational Medicine cassettes loaded with 25-mm gelatin filters were used to capture the inhalable microbial biomass. The total genomic DNA was extracted and used as a template for PCR amplification of the archaeal 16S rRNA gene. High concentrations of archaea were found in SCB bioaerosols, being as high as 10⁸ 16S rRNA gene copies per cubic meter of air. Construction and sequencing of 16S rRNA gene libraries revealed that all sequences were closely related to methanogenic archaea, such as Methanosphaera stadtmanae (94.7% of the archaeal biodiversity). Archaeal community profiles were compared by 16S rRNA gene denaturing gradient gel electrophoresis. This analysis showed similar fingerprints in each SCB and confirmed the predominance of methanogenic archaea in the bioaerosols. This study sheds new light on the nature of bioaerosols in SCBs and suggests that archaea are also aerosolized from pig manure.Over the last 30 years, swine production in Canada evolved from small family farms to industrial facilities. Pig producers have increased animal density, building mechanization, and confinement in order to decrease working and feeding time and to optimize space, leading to an increased contamination of air by bioaerosols.Even though the swine confinement building (SCB) environment has been studied for several years, little is known about the real concentration and nature of airborne microorganisms. Moreover, increasing confinement level in modern barns has raised bioaerosol levels, modifying the health risk of exposed workers. Thus far, using culture-dependent methods was the only strategy developed and used to describe SCB bioaerosol content and levels (6, 7). However, it is well known that culture-independent approaches are more likely to reveal the presence of microorganisms never suspected in most environments (2). In aerobiology, there are only a few reports using culture-independent methods (4, 15). Nehme et al. (20) applied molecular approaches to quantify and describe the bacterial aerosols in SCB and reported as much as 10⁸ bacteria per cubic meter of air, with significantly higher concentrations during winter, when the confinement is maximal. The data obtained were also compared to recent biodiversity studies of swine manures (13, 22, 25). Anaerobic gram-positive bacteria, being the greater part of the microbiological aerosols, appeared to originate from the swine manure. Those manure biodiversity studies revealed the presence of methanogenic archaea in hog wastes (22, 25). Since bacteria observed in the aerosols seem to originate from the manure, it is plausible that archaea from pig slurries are also aerosolized.We report here the characterization of the archaeal community of SCB bioaerosols by using cultivation-independent approaches. The phylogeny of airborne archaea was assessed using 16S rRNA gene sequences. Archaeal biodiversity profiles were determined with PCR-denaturing gradient gel electrophoresis (DGGE), and the concentration of aerosolized archaea was evaluated by real-time PCR by quantifying archaeal 16S rRNA gene copies in the air samples.     

Pyrosequencing Characterization of the Microbiota from Atlantic Intertidal Marine Sponges Reveals High Microbial Diversity and the Lack of Co-Occurrence Patterns

Sponges are ancient metazoans that host diverse and complex microbial communities. Sponge-associated microbial diversity has been studied from wide oceans across the globe, particularly in subtidal regions, but the microbial communities from intertidal sponges have remained mostly unexplored. Here we used pyrosequencing to characterize the microbial communities in 12 different co-occurring intertidal marine sponge species sampled from the Atlantic coast, revealing a total of 686 operational taxonomic units (OTUs) at 97% sequence similarity. Taxonomic assignment of 16S ribosomal RNA tag sequences estimated altogether 26 microbial groups, represented by bacterial (75.5%) and archaeal (22%) domains. Proteobacteria (43.4%) and Crenarchaeota (20.6%) were the most dominant microbial groups detected in all the 12 marine sponge species and ambient seawater. The Crenarchaeota microbes detected in three Atlantic Ocean sponges had a close similarity with Crenarchaeota from geographically separated subtidal Red Sea sponges. Our study showed that most of the microbial communities observed in sponges (73%) were also found in the surrounding ambient seawater suggesting possible environmental acquisition and/or horizontal transfer of microbes. Beyond the microbial diversity and community structure assessments (NMDS, ADONIS, ANOSIM), we explored the interactions between the microbial communities coexisting in sponges using the checkerboard score (C-score). Analyses of the microbial association pattern (co-occurrence) among intertidal sympatric sponges revealed the random association of microbes, favoring the hypothesis that the sponge-inhabiting microbes are recruited from the habitat mostly by chance or influenced by environmental factors to benefit the hosts.     

Symbiont Succession during Embryonic Development of the European Medicinal Leech,Hirudo verbana

The European medicinal leech, Hirudo verbana, harbors simple microbial communities in the digestive tract and bladder. The colonization history, infection frequency, and growth dynamics of symbionts through host embryogenesis are described using diagnostic PCR and quantitative PCR. Symbiont species displayed diversity in temporal establishment and proliferation through leech development.The hermaphroditic European medicinal leech (Hirudo spp.) is one of the most extensively examined animal models in neurophysiological, developmental, and behavioral studies (14) and has recently been used as a naturally occurring simple model for beneficial symbioses (5, 13). A fundamental question of microbial symbioses is how to determine the transmission mode of the symbionts between generations. Hirudo verbana reproduces by depositing eggs, which are surrounded by a cocoon. The cocoon is secreted from glandular cells of the parental mouth and usually contains 5 to 25 eggs. Each individual egg is encased by a self-enclosed vitelline membrane, referred to as the larval sac, and is bathed in a nutritious albumenous fluid (14). Complete embryonic development occurs within the cocoon and is composed of two distinct life stages, cryptolarva and juvenile. The cryptolarva transitions into a juvenile approximately midway into embryogenesis. The temporal acquisition of morphological attributes during embryonic development have been well described (3, 12, 16) (Fig. ​(Fig.11).Open in a separate windowFIG. 1.Paradigm of percent embryonic development (% ED) of the European medicinal leech, H. verbana, relative to the acquisition of digestive tract features. At 20 ± 1°C, 24 h is equivalent to 3.33% ED, with complete embryogenesis (spanning from cocoon deposition to the emergence of adult-like juveniles) requiring approximately 30 h. Staging scheme based on references 3 and 12. *, sampling time point; PD, days postcocoon deposition; prs, pairs; d, days. (Adapted from reference 12 with permission of John Wiley & Sons. Copyright 1998 Wiley-Liss, Inc.)The medicinal leech houses distinct microbial communities within its digestive tract and secretory bladders. Culturing and culture-independent profiling of the European medicinal leech, H. verbana, through fluorescence in situ hybridization, study of 16S rRNA gene clone libraries, and terminal restric-tion length polymorphism, revealed a simple and stable microbial community within the adult midgut (2, 4, 7, 8, 18). The gammaproteobacterium Aeromonas veronii and a member of the Bacteroidetes, Rikenella, were identified as consistent and dominant extracellular residents of the medicinal leech crop and intestinum. An early culture-based study detected a bacterium that is now considered to be A. veronii in the cocoon albumen and in young leeches after hatching (1). In previous electron microscopy work investigating the embryonic development of the bladders, intracellular bacteria were detected within the bladder wall and extracellular bacteria within the lumen (2, 16, 17). A recent study revealed that this microbiota is organized in distinct layers and is composed of the deltaproteobacterium Bdellovibrio, betaproteobacteria Comamonas and Sterolibacterium, members of the Bacteroidetes, Sphingobacterium and Niabella, and alphaproteobacterium Ochrobactrum spp. (10). Although the microbial constituents of the adult H. verbana digestive tract have been previously characterized, the succession, inoculum sizes, frequency of infection, and growth dynamics of these symbiont species during embryogenesis remain to be described.Putative functional roles for the crop/intestinum symbionts of the leech host include aiding in digestion, provisioning essential nutrients to the host, which are lacking in the blood meal (14), and preventing the establishment of foreign microbiota (1, 15). The symbionts localized in the bladders are suspected to play a role in the recycling of host metabolic waste into ammonia (10). The digestive tract symbionts may also display nutritional syntrophy, and possibly, A. veronii primes the host''s digestive tract to enable the establishment and persistence of the obligate anaerobic Rikenella-like bacterium, thereby contributing to the selection of the future microbiota (reviewed in reference 13). This paper details the microbial colonization patterns relative to H. verbana embryogenesis using a combination of species-specific diagnostic PCR and quantitative PCR (qPCR) analyses.     

Metaproteomics Analysis Reveals the Adaptation Process for the Chicken Gut Microbiota

The animal gastrointestinal tract houses a large microbial community, the gut microbiota, that confers many benefits to its host, such as protection from pathogens and provision of essential metabolites. Metagenomic approaches have defined the chicken fecal microbiota in other studies, but here, we wished to assess the correlation between the metagenome and the bacterial proteome in order to better understand the healthy chicken gut microbiota. Here, we performed high-throughput sequencing of 16S rRNA gene amplicons and metaproteomics analysis of fecal samples to determine microbial gut composition and protein expression. 16 rRNA gene sequencing analysis identified Clostridiales, Bacteroidaceae, and Lactobacillaceae species as the most abundant species in the gut. For metaproteomics analysis, peptides were generated by using the Fasp method and subsequently fractionated by strong anion exchanges. Metaproteomics analysis identified 3,673 proteins. Among the most frequently identified proteins, 380 proteins belonged to Lactobacillus spp., 155 belonged to Clostridium spp., and 66 belonged to Streptococcus spp. The most frequently identified proteins were heat shock chaperones, including 349 GroEL proteins, from many bacterial species, whereas the most abundant enzymes were pyruvate kinases, as judged by the number of peptides identified per protein (spectral counting). Gene ontology and KEGG pathway analyses revealed the functions and locations of the identified proteins. The findings of both metaproteomics and 16S rRNA sequencing analyses are discussed.     

Protein Profiling of the Medicinal Leech Salivary Gland Secretion by Proteomic Analytical Methods

Protein diversity of the high molecular weight fraction (molecular mass > 500 daltons) of salivary grand secretion of the medicinal leech Hirudo medicinalis has been demonstrated using methods of proteomic analysis. One-dimensional (1D) electrophoresis revealed the presence of more than 60 bands corresponding to molecular masses ranging from 11 to 483 kD. 2D-electrophoresis revealed more than 100 specific protein spots differing in molecular masses and pI values. SELDI-mass spectrometry analysis using the ProteinChip. System based on chromatography surfaces of strong anion or weak cation exchanger detected 45 individual compounds of molecular masses ranged from 1.964 to 66.5 kD. Comparison of SELDI-MS data with protein databases revealed eight known proteins from the medicinal leech. Other masses detected by proteomic analytical methods may be related to both modifications of known proteins and unknown biologically active components of leech saliva secretion.     

A Metagenomic Investigation of the Duodenal Microbiota Reveals Links with Obesity

Background

Few studies have tested the small intestine microbiota in humans, where most nutrient digestion and absorption occur. Here, our objective was to examine the duodenal microbiota between obese and normal volunteers using metagenomic techniques.

Methodology/Principal Findings

We tested duodenal samples from five obese and five normal volunteers using 16S rDNA V6 pyrosequencing and Illumina MiSeq deep sequencing. The predominant phyla of the duodenal microbiota were Firmicutes and Actinobacteria, whereas Bacteroidetes were absent. Obese individuals had a significant increase in anaerobic genera (p < 0.001) and a higher abundance of genes encoding Acyl-CoA dehydrogenase (p = 0.0018) compared to the control group. Obese individuals also had a reduced abundance of genes encoding sucrose phosphorylase (p = 0.015) and 1,4-alpha-glucan branching enzyme (p = 0.05). Normal weight people had significantly increased FabK (p = 0.027), and the glycerophospholipid metabolism pathway revealed the presence of phospholipase A1 only in the control group (p = 0.05).

Conclusions/Significance

The duodenal microbiota of obese individuals exhibit alterations in the fatty acid and sucrose breakdown pathways, probably induced by diet imbalance.     

A Synthetic Community Approach Reveals Plant Genotypes Affecting the Phyllosphere Microbiota

The identity of plant host genetic factors controlling the composition of the plant microbiota and the extent to which plant genes affect associated microbial populations is currently unknown. Here, we use a candidate gene approach to investigate host effects on the phyllosphere community composition and abundance. To reduce the environmental factors that might mask genetic factors, the model plant Arabidopsis thaliana was used in a gnotobiotic system and inoculated with a reduced complexity synthetic bacterial community composed of seven strains representing the most abundant phyla in the phyllosphere. From a panel of 55 plant mutants with alterations in the surface structure, cell wall, defense signaling, secondary metabolism, and pathogen recognition, a small number of single host mutations displayed an altered microbiota composition and/or abundance. Host alleles that resulted in the strongest perturbation of the microbiota relative to the wild-type were lacs2 and pec1. These mutants affect cuticle formation and led to changes in community composition and an increased bacterial abundance relative to the wild-type plants, suggesting that different bacteria can benefit from a modified cuticle to different extents. Moreover, we identified ein2, which is involved in ethylene signaling, as a host factor modulating the community''s composition. Finally, we found that different Arabidopsis accessions exhibited different communities, indicating that plant host genetic factors shape the associated microbiota, thus harboring significant potential for the identification of novel plant factors affecting the microbiota of the communities.