Microsporidia are natural intracellular parasites of the nematode Caenorhabditis elegans.
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ABSTRACT: For decades the soil nematode Caenorhabditis elegans has been an important model system for biology, but little is known about its natural ecology. Recently, C. elegans has become the focus of studies of innate immunity and several pathogens have been shown to cause lethal intestinal infections in C. elegans. However none of these pathogens has been shown to invade nematode intestinal cells, and no pathogen has been isolated from wild-caught C. elegans. Here we describe an intracellular pathogen isolated from wild-caught C. elegans that we show is a new species of microsporidia. Microsporidia comprise a large class of eukaryotic intracellular parasites that are medically and agriculturally important, but poorly understood. We show that microsporidian infection of the C. elegans intestine proceeds through distinct stages and is transmitted horizontally. Disruption of a conserved cytoskeletal structure in the intestine called the terminal web correlates with the release of microsporidian spores from infected cells, and appears to be part of a novel mechanism by which intracellular pathogens exit from infected cells. Unlike in bacterial intestinal infections, the p38 MAPK and insulin/insulin-like growth factor (IGF) signaling pathways do not appear to play substantial roles in resistance to microsporidian infection in C. elegans. We found microsporidia in multiple wild-caught isolates of Caenorhabditis nematodes from diverse geographic locations. These results indicate that microsporidia are common parasites of C. elegans in the wild. In addition, the interaction between C. elegans and its natural microsporidian parasites provides a system in which to dissect intracellular intestinal infection in vivo and insight into the diversity of pathogenic mechanisms used by intracellular microbes.
Project description:BackgroundIncreasing genetic and phenotypic differences found among natural isolates of C. elegans have encouraged researchers to explore the natural variation of this nematode species.ResultsHere we report on the identification of genomic differences between the reference strain N2 and the Hawaiian strain CB4856, one of the most genetically distant strains from N2. To identify both small- and large-scale genomic variations (GVs), we have sequenced the CB4856 genome using both Roche 454 (~400 bps single reads) and Illumina GA DNA sequencing methods (101 bps paired-end reads). Compared to previously described variants (available in WormBase), our effort uncovered twice as many single nucleotide variants (SNVs) and increased the number of small InDels almost 20-fold. Moreover, we identified and validated large insertions, most of which range from 150 bps to 1.2 kb in length in the CB4856 strain. Identified GVs had a widespread impact on protein-coding sequences, including 585 single-copy genes that have associated severe phenotypes of reduced viability in RNAi and genetics studies. Sixty of these genes are homologs of human genes associated with diseases. Furthermore, our work confirms previously identified GVs associated with differences in behavioural and biological traits between the N2 and CB4856 strains.ConclusionsThe identified GVs provide a rich resource for future studies that aim to explain the genetic basis for other trait differences between the N2 and CB4856 strains.
Project description:Durotaxis is a process where cells are able to sense the stiffness of substrates and preferentially migrate toward stiffer regions. Here, we show that the 1-mm-long nematode, Caenorhabditis elegans are also able to detect the rigidity of underlying substrates and always migrate to regions of higher stiffness. Our results indicate that C. elegans are able to judiciously make a decision to stay on stiffer regions. We found that the, undulation frequency, and wavelength of worms, crawling on surfaces show nonmonotonic behavior with increasing stiffness. A number of control experiments were also conducted to verify whether C. elegans are really able to detect the rigidity of substrates or whether the migration to stiffer regions is due to other factors already reported in the literature. As it is known that bacteria and other single-celled organisms exhibit durotaxis toward stiffer surfaces, we conjecture that durotaxis in C. elegans may be one of the strategies developed to improve their chances of locating food.
Project description:Microsporidia are fungi-related intracellular pathogens that may infect virtually all animals, but are poorly understood. The nematode Caenorhabditis elegans has recently become a model host for studying microsporidia through the identification of its natural microsporidian pathogen Nematocida parisii. However, it was unclear how widespread and diverse microsporidia infections are in C. elegans or other related nematodes in the wild. Here we describe the isolation and culture of 47 nematodes with microsporidian infections. N. parisii is found to be the most common microsporidia infecting C. elegans in the wild. In addition, we further describe and name six new species in the Nematocida genus. Our sampling and phylogenetic analysis further identify two subclades that are genetically distinct from Nematocida, and we name them Enteropsectra and Pancytospora. Interestingly, unlike Nematocida, these two genera belong to the main clade of microsporidia that includes human pathogens. All of these microsporidia are horizontally transmitted and most specifically infect intestinal cells, except Pancytospora epiphaga that replicates mostly in the epidermis of its Caenorhabditis host. At the subcellular level in the infected host cell, spores of the novel genus Enteropsectra show a characteristic apical distribution and exit via budding off of the plasma membrane, instead of exiting via exocytosis as spores of Nematocida. Host specificity is broad for some microsporidia, narrow for others: indeed, some microsporidia can infect Oscheius tipulae but not its sister species Oscheius sp. 3, and conversely some microsporidia found infecting Oscheius sp. 3 do not infect O. tipulae. We also show that N. ausubeli fails to strongly induce in C. elegans the transcription of genes that are induced by other Nematocida species, suggesting it has evolved mechanisms to prevent induction of this host response. Altogether, these newly isolated species illustrate the diversity and ubiquity of microsporidian infections in nematodes, and provide a rich resource to investigate host-parasite coevolution in tractable nematode hosts.
Project description:Animal predators can track prey using their keen sense of smell. The bacteriovorous nematode Caenorhabditis elegans employs sensitive olfactory sensory neurons that express vertebrate-like odor receptors to locate bacteria. C. elegans displays odor-related behaviors such as attraction, aversion and adaptation, but the ecological significance of these behaviors is not known. Using a combination of food microbiology and genetics, we elucidate a possible predator-prey relationship between C. elegans and lactic acid bacteria (LAB) in rotting citrus fruit. LAB produces the volatile odor diacetyl as an oxidized by-product of fermentation in the presence of citrate. We show that C. elegans is attracted to LAB when grown on citrate media or Citrus medica L, commonly known as yuzu, a citrus fruit native to East Asia, and this attraction is mediated by the diacetyl odor receptor, ODR-10. We isolated a wild LAB strain and a wild C. elegans-related nematode from rotten yuzu, and demonstrate that the wild nematode was attracted to the diacetyl produced by LAB. These results not only identify an ecological function for a C. elegans olfactory behavior, but contribute to the growing understanding of ecological relationships between the microbial and metazoan worlds.
Project description:Microsporidia are fungi-like parasites that have the smallest known eukaryotic genome, and for that reason they are used as a model to study the phenomenon of genome decay in parasitic forms of life. Similar to other intracellular parasites that reproduce asexually in an environment with alleviated natural selection, Microsporidia experience continuous genome decay that is driven by Muller's ratchet-an evolutionary process of irreversible accumulation of deleterious mutations that lead to gene loss and the miniaturization of cellular components. Particularly, Microsporidia have remarkably small ribosomes in which the rRNA is reduced to the minimal enzymatic core. In this study, we analyzed microsporidian ribosomes to study an apparent impact of Muller's ratchet on structure of RNA and protein molecules in parasitic forms of life. Through mass spectrometry of microsporidian proteome and analysis of microsporidian genomes, we found that massive rRNA reduction in microsporidian ribosomes appears to annihilate the binding sites for ribosomal proteins eL8, eL27, and eS31, suggesting that these proteins are no longer bound to the ribosome in microsporidian species. We then provided an evidence that protein eS31 is retained in Microsporidia due to its non-ribosomal function in ubiquitin biogenesis. Our study illustrates that, while Microsporidia carry the same set of ribosomal proteins as non-parasitic eukaryotes, some ribosomal proteins are no longer participating in protein synthesis in Microsporidia and they are preserved from genome decay by having extra-ribosomal functions. More generally, our study shows that many components of parasitic cells, which are identified by automated annotation of pathogenic genomes, may lack part of their biological functions due to continuous genome decay.
Project description:The acquisition of genes by horizontal transfer can impart entirely new biological functions and provide an important route to major evolutionary innovation. Here we have used ancient gene reconstruction and functional assays to investigate the impact of a single horizontally transferred nucleotide transporter into the common ancestor of the Microsporidia, a major radiation of intracellular parasites of animals and humans. We show that this transporter provided early microsporidians with the ability to steal host ATP and to become energy parasites. Gene duplication enabled the diversification of nucleotide transporter function to transport new substrates, including GTP and NAD+, and to evolve the proton-energized net import of nucleotides for nucleic acid biosynthesis, growth and replication. These innovations have allowed the loss of pathways for mitochondrial and cytosolic energy generation and nucleotide biosynthesis that are otherwise essential for free-living eukaryotes, resulting in the highly unusual and reduced cells and genomes of contemporary Microsporidia.
Project description:BackgroundThe nematode Caenorhabditis elegans was the first multicellular organism to have its genome fully sequenced. Over the last 10 years since the original publication in 1998, the C. elegans genome has been scrutinized and the last gaps were filled in November 2002, which present a unique opportunity for examining genome-wide segmental duplications.ResultsHere, we performed analysis of the C. elegans genome in search for segmental duplications using a new tool -- OrthoCluster -- we have recently developed. We detected 3,484 duplicated segments -- duplicons -- ranging in size from 234 bp to 108 Kb. The largest pair of duplicons, 108 kb in length located on the left arm of Chromosome V, was further characterized. They are nearly identical at the DNA level (99.7% identity) and each duplicon contains 26 putative protein coding genes. Genotyping of 76 wild-type strains obtained from different labs in the C. elegans community revealed that not all strains contain this duplication. In fact, only 29 strains carry this large segmental duplication, suggesting a very recent duplication event in the C. elegans genome.ConclusionThis report represents the first demonstration that the C. elegans laboratory wild-type N2 strains has acquired large-scale differences.
Project description:Genetic variation in host populations may lead to differential viral susceptibilities. Here, we investigate the role of natural genetic variation in the Intracellular Pathogen Response (IPR), an important antiviral pathway in the model organism Caenorhabditis elegans against Orsay virus (OrV). The IPR involves transcriptional activity of 80 genes including the pals-genes. We examine the genetic variation in the pals-family for traces of selection and explore the molecular and phenotypic effects of having distinct pals-gene alleles. Genetic analysis of 330 global C. elegans strains reveals that genetic diversity within the IPR-related pals-genes can be categorized in a few haplotypes worldwide. Importantly, two key IPR regulators, pals-22 and pals-25, are in a genomic region carrying signatures of balancing selection, suggesting that different evolutionary strategies exist in IPR regulation. We infected eleven C. elegans strains that represent three distinct pals-22 pals-25 haplotypes with Orsay virus to determine their susceptibility. For two of these strains, N2 and CB4856, the transcriptional response to infection was also measured. The results indicate that pals-22 pals-25 haplotype shapes the defense against OrV and host genetic variation can result in constitutive activation of IPR genes. Our work presents evidence for balancing genetic selection of immunity genes in C. elegans and provides a novel perspective on the functional diversity that can develop within a main antiviral response in natural host populations.
Project description:Bio-electrospray, the direct jet-based cell handling apporach, is able to handle a wide range of cells. Studies at the genomic, genetic, and the physiological level have shown that, post-treatment, cellular integrity is unperturbed and a high percentage (>70%, compared to control) of cells remain viable. Although, these results are impressive, it may be argued that cell based systems are oversimplistic. This study utilizing a well characterised multicellular model organism, the non-parasitic nematode Caenorhabditis elegans. Nematodes were subjected to bio-electrosprays to demonstrate that bio-electrosprays can be safely applied to nematodes.
Project description:The growth of pathogens is dictated by their interactions with the host environment1. Obligate intracellular pathogens undergo several cellular decisions as they progress through their life cycles inside host cells2. We have studied this process for microsporidian species in the genus Nematocida as they grew and developed inside their co-evolved animal host, Caenorhabditis elegans3-5. We found that microsporidia can restructure multicellular host tissues into a single contiguous multinucleate cell. In particular, we found that all three Nematocida species we studied were able to spread across the cells of C. elegans tissues before forming spores, with two species causing syncytial formation in the intestine and one species causing syncytial formation in the muscle. We also found that the decision to switch from replication to differentiation in Nematocida parisii was altered by the density of infection, suggesting that environmental cues influence the dynamics of the pathogen life cycle. These findings show how microsporidia can maximize the use of host space for growth and that environmental cues in the host can regulate a developmental switch in the pathogen.