-
[
International Worm Meeting,
2015]
Like us, C. elegans lives in a microbial world. In its natural habitats of rotting fruits and vegetation, C. elegans encounters a diverse array of microbes, where they serve as diet, microbiota or pathogens. C. elegans is highly tuned and equipped to sense and response to the milieu of microbial products (xenobiotics) that they are bombarded with, as they navigate this microbial landscape. Indeed, several recent studies have demonstrated that animals monitor basic cellular subsystems for microbial targeting (presumably through decreased functionality/efficiency or the like), though the extent to which beneficial microbes may interface with these systems is unknown. Small RNA pathways play a central role in regulating many of the transcriptional and developmental programs that are responsive to microbes, in addition to directly mediating anti-viral immunity. Thus, this study takes a broad look at natural microbes that may specifically engage small RNA pathways to regulate C. elegans physiology.To examine this question, we screened a panel of 565 microbes ('BIGb collection') isolated from C. elegans' natural habitats for microbial-enhancers or -reducers of RNAi (mERI or mRDE, respectively) via co-feeding with several RNAi clones in E. coli (somatic and germline) or alone with a panel of transgenic reporters. In co-feeding experiments, nearly 20% of the microbes reduced RNAi effectiveness (113), while 7.5% enhanced RNAi (42). There was also a strong correlation between the impact of a microbe on C. elegans physiology-beneficial (faster growth, unstressed) or detrimental (slow growth/death, activation of stress reporters)-and its impact on RNAi pathways, with mRDE isolates being more likely to be detrimental and vice versa. Since co-feeding can affect uptake of dsRNA-producing E. coli, we also validated these microbes for impacts on RNAi using a panel of transgene-silencing reporter strains. Together, these data implicate a number of often pathogens (e.g., Pseudomonas, Stenotrophomonas and Enterobacter) as mRDE and beneficial microbes (e.g., Providencia and Bacillus) as mERI. In parallel, we also demonstrate that microbial RNA directly engages a subset of RNAi pathways in regulation of C. elegans growth. Together, these studies expand our understanding of the host systems that are under microbial influence and regulate host health. .
-
Rowedder, Holli, Kirienko, Natalia, Ruvkun, Gary, Breen, Peter, Zhang, Xinrui, Qi, Yan
[
International Worm Meeting,
2013]
A wide range of bacteria and fungi produce potent xenobiotics that inhibit the growth of other organisms in ecological conflict. In defense, the target organisms have evolved extensive countermeasures to detect and detoxify the foreign toxins, as well as to repair the damage caused by the insults. The ribosome represents a common target of many natural toxins of diverse chemical nature. For example, a small-molecule drug hygromycin produced by the bacterium Streptomyces hygroscopicus and a protein toxin exotoxin A produced by the human bacterial pathogen Pseudomonas aeruginosa can cause ribotoxic stress in C. elegans by inhibiting the translation elongation step of the worm ribosome. Using hygromycin-induced developmental arrest as readout, a genome-wide RNAi screen was carried out to identify genes that regulate the ribotoxic stress response. We show that many hits from this screen are also involved in defense against Pseudomonas infection in worms. Microarray and small RNA deep-sequencing experiments were performed to profile the mRNA and small RNA expression levels under ribotoxic stress. We find that many innate immunity, detoxification and metabolism genes are differentially regulated in worms under ribotoxic stress, and the level of differential regulation of these genes is attenuated by inactivation of the genes identified in the hygromycin RNAi screen. Our findings suggest a novel model of xenobiotic detection and detoxification, in which decrement in the essential cellular functions serves as a general signal of xenobiotic stress. Understanding the genetic network behind xenobiotic detection, detoxification, and surveillance of essential cellular functions will provide insights into human response to medicines, drug resistance, and stress response.
-
[
International Worm Meeting,
2013]
Like us, C. elegans lives in a microbial world. In its natural habitats of rotting fruits and vegetation, these nematodes proliferate as they dine on an array of microbes. Interactions with microbes span a spectrum from constant confrontation (pathogens) to relative indifference (food) to perhaps even mutual benefit (symbionts). This study identifies these natural microbes and addresses whether microbiome composition influences proliferation of C. elegans in the wild.
To examine this question, we sequenced bacterial 16S (SSU) rDNA amplicons from habitats with wild C. elegans populations collected in France and Spain. Our results show that C. elegans encounters a broad array of bacteria in the wild-especially the divisions (phyla) of Proteobacteria, Bacteroidetes, Firmicutes and Actinobacteria. An abundance-weighted comparisons of phylogenetic differences (UniFrac) showed distinct clustering by habitat type, as rotting apples clustered separately from other habitats sequenced. Further, rotting apples clustered by large presence of proliferating or small non-proliferating (dauer) populations of worms. C. elegans appear to proliferate in apples with 'simpler' microbiomes (lower diversity, fewer species and Proteobacteria-rich). Specific alpha-proteobacteria were particularly enriched in apples with proliferating worms, while a number of genera were consistently found in apples with non-proliferating worms (e.g., Pseudomonas, several Bacteroidetes, etc.). Population size also correlated with apple rottenness, suggesting bacterial load is key to growth as well.
Similarly, Proteobacteria content does affect C. elegans (N2) growth rate in the lab, as worms grew faster on mixtures (and single isolates) with 80% Proteobacteria versus those with 40% Proteobacteria. Together, these studies define the microbial diet of C. elegans and implicate the natural microbiome as a key determinant of C. elegans' growth in the wild.
-
Carr, Chris, Rowedder, Holli, Plunkett, Guy, Melo, Justine, Durfee, Tim, Nusbaum, Chad, Glasner, Jeremy, Russ, Carsten, Ruvkun, Gary, Sykes, Sean, Samuel, Buck S., Young, Sarah
[
International Worm Meeting,
2011]
Like other metazoans, C. elegans fitness (success) within its microbe-rich habitats depends on a tight balance of energy acquisition and expenditure. Thus, it is also highly tuned to microbial cues that allow it to separate potential food or friend from foe. Accordingly, some microbial signals have been postulated to influence fat storage in parallel to endogenous endocrine cues. Several studies also show that the E. coli-adapted N2-Bristol strain is especially sensitive to 'minor' differences in E. coli strains: faster growth rates, increased progeny delivery rates, and less fat retention is seen when worms consume HB101 compared to OP50. Perhaps due to this fitness benefit, worms also exhibit increased satiety and a behavioral preference for HB101. Thus, we have sought to identify the E. coli gene products that modulate C. elegans fitness. To this end, we have sequenced E. coli genomes routinely used in C. elegans cultivation: HB101 (2 isolates), OP50 (2 isolates) and HT115. Despite little variation among strain isolates, 350 and 412 genes are 'unique' to OP50 and HB101, respectively. Many are organized into clusters, and represent a range of gene functions: e.g., carbohydrate utilization (96), cell wall/LPS modification (42), amino acid metabolism (21), regulation (41), the Cascade system (6) and fatty acid metabolism (5). Phenotype microarrays were also used to confirm the metabolic defects. In order to systematically test the impact of these microbial gene products on C. elegans' fitness, we assembled nearly 200 single gene mutants with defined function in a 'neutral' and consistent genetic background (E. coli K12). We then used a number of assays to test a mutant's impact on N2 growth, broods, body size and fat storage. Our analyses indicate that both genes in core metabolism and transport/biosynthesis of conserved mediators of host interaction-autoinducers, biogenic amines, short-chain fatty acids and LPS-influence N2 fitness. Studies of these small molecules as sensory or nutritive cues to C. elegans directly or via regulation of E. coli metabolism are ongoing. However, results so far indicate that the microbial milieu of signals may be just as important of a determinant of C. elegans' fitness as the nutritional potential for supporting growth of a population within a given habitat.