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[
Evolutionary Biology of Caenorhabditis and Other Nematodes,
2010]
Caenorhabditis briggsae is a useful species with which to pursue evolutionary and genomic questions by comparison with C. elegans. However, in C. briggsae the genetic maps and genome assembly necessary for robustly conducting experiments, such as mapping mutations, are not of comparable quality to those available for C. elegans. To improve the genetic map and genome assembly of C. briggsae, 180 AF16xHK104 and HK104xAF16 advanced-intercross recombinant inbred lines (AI-RIL) were genotyped at 1,536 single nucleotide polymorphism (SNP) markers using the Illumina GoldenGate platform. Following quality control, the final dataset comprised 167 AI-RIL typed at 1,034 markers. Employing AI-RIL increased the number and decreased the size of haplotype blocks per chromosome, and the dense panel of genetic markers allowed most previously unplaced sequence contigs to be ordered within chromosome assemblies. Additional issues with the present genome assembly, including contig misassignment to chromosomes, have been resolved to produce a new genome assembly (
cb4). Our genotyping data also reveal interesting population genetic phenomena. Analysis of parental allele fraction of the AI-RIL reveals strong bias of three autosomes in favor of the HK104 parental allele. The bias of one autosome is independent of cross direction and due to a slow-growth phenotype elicited in a hybrid genome. The bias of the other two autosomes is strongly dependent on cross direction. Comparison of inter-chromosomal linkage disequilibrium (LD) in the AI-RIL also reveals the presence of cross-direction-specific LD. Together, the genome-wide study of marker segregation in C. briggsae AI-RIL reveals evidence of inter-strain genomic incompatibilities that suggest the onset of genomic divergence and potentially incipient speciation. Efforts are currently under way to identify the incompatibility loci involved.
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[
International Worm Meeting,
2019]
WormBase (www.wormbase.org) has been serving the scientific community for 19 years as a central repository of genomic and genetic information for C. elegans and other nematodes. We continually enlarge and enrich the included data, develop tools and displays for exploring those data on the website, and improve the back-end database and infrastructure to allow us to capture more data and serve it faster. For example, we are now up-to-date on protein-protein interactions and are developing new displays for these data, and ParaSite now has over 100 nematode genomes! We are fully engaged in the Alliance of Genome Resources (www.alliancegenome.org), which uses the combined expertise of seven information resources to deliver better services to all our communities: advances at WormBase such as our AI-generated descriptions of gene function are now used across the Alliance while advances at the Alliance such as Gene Ontology (GO) ribbons that concisely summarize annotations or enrichment of human disease model annotations are now visible on www.wormbase.org. We are using AI to make the best use of your time: after publication, we have been emailing authors to help us extract information. We are now using AI on a revised Author First Pass form for authors to confirm rather than enter data in their papers, thereby saving keystrokes. Our experiment in making the publication process knowledge-base compatible, www.microPublication.org has taken on a life of its own. Our curators have started to visit various universities and regional meetings to give on-site tutorials and get direct feedback; please contact us if you are interested.
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Jorgensen, C, Chahal, G, Haji Adineh, S, Ortega, B, Aguilar, A, Ross, J
[
International Worm Meeting,
2019]
Although mitochondria are usually maternally inherited, substantial evidence from multiple taxa reveals that hybridization often promotes transmission of sperm-borne mitochondrial into the fertilized oocyte. Given prior reports of such paternal "leakage" of mitochondria in intra-species C. briggsae crosses, we conducted new experiments to refine our understanding of the genetic basis and tempo of paternal mitochondrial transmission (PMT). Analysis of paternal mitochondrial haplotypes in replicate hybrid lines revealed persistent but transient heteroplasmy that never persisted beyond ten generations. Thus, PMT appears to occur regularly following hybridization, but heteroplasmy is usually resolved in favor of the maternal mitotype. Additionally, no paternal mitotypes were detected among a sample of existing C. briggsae advanced-intercross recombinant inbred lines (AI-RIL), which could suggest that using recombination to disrupt a potentially complex genetic mechanism that regularly acts to prevent paternal transmission does not result in sustained homoplasmy of the paternal mitotype. However, crosses involving AI-RIL pseudofemales do occasionally result in PMT, further implicating the role of a hybrid maternal nuclear genome in facilitating PMT. Current efforts in C. elegans are testing the hypothesis that PMT is dependent on somatic sex, such that hermaphrodites do not perform paternal mitochondrial elimination during self-fertilization but do eliminate non-self sperm mitochondria when mated with a male.
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[
International Worm Meeting,
2013]
Bacterial group behaviors are governed by a process called quorum sensing, in which bacteria produce, secrete, and detect extracellular signal molecules called autoinducers (AIs). Vibrios produce multiple AIs, some enable intra-species communication and others that promote inter-species communication. Vibrio cholerae produces an intra-species AI called CAI-1 that is a 13 carbon long fatty acyl molecule and the interspecies signal called AI-2 that is a boron-containing furanone. The information contained in the AIs is funneled into a shared phosphorelay signaling cascade that controls virulence, biofilm formation, and other traits. The bacteriovorous nematode, Caenorhabditis elegans, also uses small molecules to interpret its environment. A class of C. elegans-derived molecules called ascarosides influence nematode behaviors including attraction, repulsion, and mating. The presence of bacteria stimulates chemotaxis, egg-laying, and feeding in C. elegans, however, the bacteria-produced molecules that the nematode detects to control these phenotypes are largely unknown. We demonstrate that in addition to playing a vital role in quorum-sensing-regulated behaviors in V. cholerae, CAI-1 also influences behavior in C. elegans. C. elegans is more strongly attracted to V. cholerae than to its food source E. coli HB101 and C. elegans prefers V. cholerae that produces CAI-1 over a V. cholerae mutant for CAI-1 production. Consistent with this finding, robust chemoattraction occurs to synthetic CAI-1. CAI-1 is detected by the sensory neuron AWCON. Laser ablation of this cell, but not other amphid sensory neurons, abolished chemoattraction to CAI-1. To define which moieties of CAI-1 are crucial for recognition by C. elegans, we synthesized CAI-1 analogs and tested whether they promote chemoattraction. The fatty-acid chain length as and the precise position of the CAI-1 ketone group are the key features required for mediating CAI-1-directed nematode behavior. Together, these analyses define a bacteria-produced signal and the nematode detection apparatus that permit interkingdom communication.
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[
International Worm Meeting,
2009]
Vibrio cholerae (VC), the causative agent of cholera in humans, is responsible for devastating epidemics and pandemics across the world. The major virulence factor underlying the pathogenesis of cholera is cholera toxin (CT). However, CT negative VC non-O1 and non-O139 strains and CT deleted vaccine mutant strains are still capable of causing disease symptoms through mechanisms that are currently unclear. Vibrio cholerae cause lethality, growth retardation and escape behavior in Caenorhabditis elegans via cholera toxin (CT), and toxin co-regulated pili (TCP) independent process (1, 2). Absence of the CT and TCP response in C. elegans model may help to reveal the role of other toxins of VC that might otherwise be masked by these major virulence factors. CVD110, a V.cholerae vaccine strain, lacking several virulence factors such as zonula occludens toxin (zot), accessory toxin (ace), hemolysin (hly A) and cholera toxin A subunit gene (ctx A), showed attenuated killing in C. elegans (3,4). We are conducting microarray experiments to define host immune response genes expressed upon exposure to VC virulence factors. Differential expression profiling of C. elegans exposed to wild type VC versus CVD110 are being done using Affymetrix expression microarrays. Results of these experiments will be presented. (1) Vaitkevicius K. et al. PNAS, 103 (2006) 9280-9285 (2) Cinar HN. et al. 16 th International C. elegans Meeting, 2007 (3) Michalski J. et al. Infection and Immunity 61 (1993) 4462-4468 (4) Cinar HN. et al. Aging Stress and Pathogenesis Meeting, 2008.
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[
International Worm Meeting,
2015]
The Dobzhansky-Bateson-Muller model suggests that neutral genetic differences unique to one of two populations can lead to a reduction in hybrid fitness (hybrid dysfunction) when individuals from those two populations hybridize, combining those alleles. Additional mutations also contribute to hybrid dysfunction, pushing the two populations along the continuum to becoming distinct species. Thus, although it is easy to identify different species, it is difficult to determine the identity and order of genetic events that establish hybrid dysfunction and species formation. One way to circumvent this issue is to study cases in which hybrids are only slightly less fit than their parents, a situation that might represent the initiation of speciation. Such a situation exists in inter-population hybrids of Caenorhabditis briggsae. Two populations, AF16 and HK104, can be hybridized, but two distinct dysfunction phenotypes have been identified in their hybrids. In the first (Ross et al. 2011 PLoS Genetics; Baird and Stonesifer 2012 Worm), a small percent of F2 hybrids exhibit developmental delay, taking longer to reach adulthood than their wild-type siblings. In the second, genetic evidence from AF16-HK104 advanced-intercross recombinant inbred lines (AI-RIL) also suggested the presence of negative mitochondrial-nuclear epistatic interactions that might reduce hybrid fitness (Ross et al. 2011 PLoS Genetics). We present evidence from F10 mitochondrial-nuclear hybrids that a range of dysfunction phenotypes (including mitochondrial physiology and life history traits) are produced by exchanging the mitochondrial genome of one strain for the other, confirming the existence of mito-nuclear incompatibility. We also unexpectedly uncovered empirical evidence of occasional male mitochondrial transmission, challenging the common knowledge that mitochondria are maternally inherited and raising the possibility of using C. briggsae to study the cellular mechanisms that normally act to prevent such inheritance. Finally, three independent mapping approaches, involving F2 bulk segregant mapping, AI-RIL mapping, and near-isogenic line (NIL) mapping have begun to narrow down the region of C. briggsae chromosome III that contains one of the epistatic loci contributing to hybrid developmental delay. In sum, we present multiple lines of evidence that suggest the genetic divergence between AF16 and HK104 has placed these populations on the speciation continuum. Ongoing efforts seek to identify the molecular causes for these hybrid dysfunction phenotypes to pinpoint the genetic basis of species formation.
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[
International C. elegans Meeting,
1991]
Mut-5 is an endogenous mutator of Caenorhabditis elegans, causing Tc1 transposition and excision in the germline. Mut-5 was first mapped on chromosome 11 between
dpy-10 and
rol-1 (Mori et ai, 1988). Subsequent experiments showed that
mut-5 is located between
dpy-10 and
unc-4, to the left of
vab-9, on chromosome 11. In order to fine-map and identify
mut-5 a three factor cross between RP3 (
mut-5(
st701) unc- 4
(e120) 11:
unc-22(
st136::Tc1) IV) and RP5 (
dpy-10(
e128)
vab-9(
e1744) 11) was performed and a total of 12 Vab-9 Unc-22 recombinants was obtained. Mutator activity of the recombinants was tested by scoring for reversion of the
unc-22(
st136::Tc1) allele. 5 Vab-9 Unc-22 recombinants proved to be mutator positive and 7 mutator negative, which mapped
mut-5 near
zyg-11 on chromosome 11. Because the mutator is assumed to be a mobile element, the Vab-9 Unc22 recombinants were tested for polymorphisms cosegregating with
mut-5. Therefore chromosomal DNA of 9 Vab-9 Unc-22 recombinants (3 mutator positive and 6 mutator negative) was digested with EcoRI and analyzed in Southern blotting experiments. One of us (IM) identified 3 Tc1 elements cosegregating with
mut-5, Tc1#40, Tc1#55 and Tc1#118: these mapped in the area where
mut-5 was mapped by three factor crosses. Those elements were cloned and sequenced (IM) and their flanking sequences were used to investigate whether one of the Tc1 elements cosegregated with
mut-5 in the recombinants of the
dpy-10-
vab-9 area. Indeed one of them, Tc1#40, was present in all tested mutator positive recombinants and absent in the mutator negative recombinants. These results show that Tc1#40 maps in close proximity of
mut-5. We consider it likely that
mut-5 is Tc1#40. Since the sequence of this element shows no consistent differences with other Tc1 elements, Tc1#40 cannot be a mutator as a result of its sequence per se. Possibly the chromosomal position of Tc1#40 causes transcription of the element at the proper time and place for germline activation of Tc1. This hypothesis could also provide an explanation for the fact that Tc1 excises somatically in a non-mutator strain (Emmons et ai, 1983): copies of Tc1 could be transcribed in the soma but not in the germline. We are currently investigating this hypothesis by generating animals transgenic for Tc1#40 and flanking C. elegans sequences
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[
International C. elegans Meeting,
2001]
To build upon knowledge gained from the genome of C. elegans , we have begun generating Expressed Sequence Tags (ESTs) from parasitic (and free-living) nematodes. This project will generate >225,000 5' ESTs from 14 species by 2003. Additionally, the Sanger Centre and Edinburgh Univ. will complete 80,000 ESTs from 7 species. Through these combined efforts, we anticipate the identification of >80,000 new nematode genes. At the GSC, approximately 35,000 ESTs have been generated to date including sequences from Ancylostoma caninum, Heterodera glycines, Meloidogyne incognita and javanica, Parastrongyloides trichosuri, Pristionchus pacificus, Strongyloides stercoralis and ratti, Trichinella spiralis, and Zeldia punctata . We will report on our progress in sequence analysis, including the creation of the NemaGene gene index for each species by EST clustering and consensus sequence generation, identification of common and rare transcripts, and identification of genes with orthologues in C. elegans and other nematodes. All sequences are publicly available at www.ncbi.nlm.nih.gov/dbEST. NemaGene sequences and project details are available at WWW.NEMATODE.NET. We would like to thank collaborators who have provided materials and ideas for this project including Prema Arasu, David Bird, Rick Davis, Warwick Grant, John Hawdon, Doug Jasmer, Andrew Kloek, Thomas Nutman, Charlie Opperman, Alan Scott, Ralf Sommer, and Mark Viney. This work is funded by NIH-AI-46593, NSF-0077503, and a Merck / Helen Hay Whitney Foundation fellowship.
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[
International Worm Meeting,
2005]
Codon usage has direct utility in molecular characterization of a species and also serves as a marker for molecular evolution. In order to better understand codon usage within the diverse phylum Nematoda, we analyzed a total of 265,494 ESTs in 93,645 clusters from 30 nematode species. Putative translations, obtainable for 75% of clusters, were based upon homology to known or predicted proteins. The full genomes of Caenorhabditis elegans and C. briggsae were also examined. A total of 25,871,325 codons were analyzed and a definitive codon usage table for all species was generated. Similarity in codon usage can be quantified by the chi-square statistic. Related nematodes have previously been observed to have similar codon usage but the evolutionary distances at which conservation diminishes had not been established. We show that codon usage similarity in Nematoda is a short-range phenomenon, generally persisting over the breadth of a genus but then rapidly diminishing within each clade. A second focus was the underlying factors that bias codon usage. In comparing species, we find a strong correlation between the overall AT/GC content of the genome and similarity in codon usage. Surprising, differences exist among species and clades in the degree of codon-usage bias as measured by effective number of codons (ENC), indicating potential differences in selective pressures or population dynamics. This study was supported by NIH-NIAID AI-46593.
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[
International C. elegans Meeting,
2001]
Two web sites have been established to allow easier access to nematode sequences from species other than C. elegans and C. briggsae ; WWW.NEMATODE.NET is maintained by the Genome Sequencing Center (GSC) at Washington University in collaboration with North Carolina State University, and WWW.NEMATODES.ORG is maintained by Mark Blaxter's lab at the University of Edinburgh. Useful features being built for NEMATODE.NET include the following - 1) Searches: All nematode expressed sequence tags (ESTs) generated at the GSC, currently 32,000 from 10 species, and NemaGene clusters built from these ESTs, are available for BLAST and text searching. Searches can be directed by species, library, or nematode clade in a way that is not possible using the NCBI EST database dbEST. 2) FTP: All EST project data can be downloaded for local analysis including FASTA files and sequence trace image files. 3) Trace Viewer: Fluorescent trace representations for each EST can be viewed. Traces can sometimes provide additional sequence information not included in the EST due to quality value cut-offs. 4) Project Updates: Information is available about libraries in construction and sequencing in progress as the project expands toward 235,000 ESTs. 5) Clone Requests: Details on clone availability and ordering procedure are provided. 6) Links: The site includes an up-to-date set of 300 links to information on human, animal, and plant parasitic nematodes. Further plans for NEMATODE.NET include linking of ESTs to their closest C. elegans homologues by DAS third-party curation of Wormbase. This work is funded by NIH-AI-46593, NSF-0077503, and a Merck / Helen Hay Whitney Foundation fellowship.